Geopolitics in the Middle East Essay Example | Topics and Well Written Essays - 750 words

Geopolitics in the Middle East - Essay Example Despite the struggles of the advocates of change, with the neo-conservatives envisioning an end for the, sort of chaotic Middle East society, the fight leads to a different scenario than what was expected. Peace is still so elusive, terrorist threats still exist, and instability is still greatly felt. We examined the reasons behind the pitfalls of this political blitzkrieg based on the articles written by Jeffrey Goldberg entitled "After Iraq" and the article "Parties of God: The Bush doctrine and the rise of Islamic Democracy" written by Ken Silverstein. Goldberg generally expressed the fact that the political agenda of the US and its allies are contrary to what Iraq's minority groups and even the rest of the countries in the region are looking for. Silverstein in the other hand expressed the fact that the Bush doctrine and the western democratic system do not fit the Islamic cultures and ways. While the US and the rest of its allies are eyeing to push their own vested interests in the Middle East campaign, the Middle Eastern nations affected by the current transitions in the other hand are looking at it in a different perspective. The war on Iraq was generally aimed at ousting Saddam Hussein, a dictatorial leader, for the major reason of removing a danger to social equality in the region, which Hussein had proven to be a threat, and instate a unified democratic government. . The plans may have been understood plainly by both the allies and Arab compatriots, like the Kurds, Turkey, Egypt, Jordan, and the rest, and laid open for them to work through, but beneath the shadows of their support, the Middle eastern countries, and minority regions within Iraq, are hoping to push through with their own agenda, for their own people, for their leader's interest. Take for example the case of the Kurds. In a personal interview by Jeffrey Goldberg with Abdul Rahman Mustafa, the Kurdish-Iraqi governor of Kirkuk in northern Iraq, considered as domains of the Kurds, it appeared that the Kurds really wanted independence from Iraq (Goldberg 79). The article of Goldberg speak further of other regions in the middle east that wanted to be governed based on their own cultural minority principles such as the "Shiites did not want to be ruled from Baghdad by a Sunni minority"(78), and other minority groups from Egypt, Israel, and other areas want their own sovereignty. In other words, the zeroing in was just superficial. The question is why The answer perhaps lies on the cultural differences between the east and the west, and the way these two civilizations are molded through time. We agree on Goldberg's idea that western democratic strategies don't seem to fit with the Middle Eastern cultures, which are highly diverse. Islam, which is the main religion in the Middle Eastern countries, is a diverse religion with differing principles across different minorities. While the west is generally Christian, although differing in minor teachings, have common core values that has high regard to peace and order. There is no Christian jihad, to cite a particular difference, while the Muslim jihad is primarily exploited by Muslim extremists and used to push their ideals and principles. In the western world, as a common knowledge, governance in civic

Human Carbonic Anhydrase II

Human Carbonic Anhydrase II Human carbonic anhydrase II is one of the fastest studied enzymes known with a variety of roles in reaction catalysis. Its primary function is to catalyze the reversible hydration reaction of carbon dioxide. In addition to carbon dioxide hydration, it is also capable of other latent skills, such as catalyzing esterase activity. The ability of human carbonic anhydrase II to function as a catalyst derives from key residues in and around the active site that play crucial roles in the mechanism. Substitutions to two of those particular key amino acids were performed via Quick-change site directed mutagenesis: H64A and V142D, to investigate the particular role they have in the catalytic active site. Various kinetic experiments and structural analyses were performed on wild-type carbonic anhydrase and the mutants to discern and compare their activity to each other and to literature, including Michaelis-Menten parameters for PNPA hydrolysis, CO2 hydration, and inferring function molecular m odelling. Though the same trends can be seen as the literature, individual values were found to be much lower owing to errors in measurement and equipment. Trends were found to coincide with the mutants known roles in the active site: His64 is the proton shuttle that facilitates proton transfer during the rate limiting step and Val142 participates in the hydrophobic pocket to bind and recruit substrates to interact with the active site. Mutations to both of these sites show that enzyme efficiency and activity strongly decreases. Introduction Human carbonic anhydrase II (hCAII) is a zinc metalloenzyme that catalyzes the following reversible reaction: . The enzyme commonly functions to help shuttle carbon dioxide in red blood cells to rid the body of metabolic waste, and catalyzes the hydrolysis of many aromatic esters [1, 2]. Structurally, a zinc ion is located in the active site, coordinated to 3 histidine residues (H94, H96, H119) and usually a hydroxide ion or water molecule [2]. The mechanism of hCAII proceeds through two major steps: 1) the conversion of carbon dioxide to bicarbonate, and 2) the regeneration of Zn-OH by proton transfer. The active hydroxide that is bound to zinc nucleophilically attacks a nearby carbon dioxide molecule, resulting in a bicarbonate ion binding to zinc [3]. The zinc-oxygen bond breaks to subsequently release a bicarbonate ion, which is replaced with water [3]. The Znà ¢Ã¢â€š ¬Ã¢â‚¬â„¢OH bond is regenerated by a proton transfer to the external buffer, which is facilitated by the His64 residue [3]. The proton transfer step is the rate limiting step of the reaction [3]. The diazole side chain on the histidine residue is what gives it the ability to be a proton acceptor and donor. Mutations in that position (His64) usually result in decreased enzyme activity due to a lack of proton transfer; however the reaction does proceed to a lesser degree without an active His residue, possibly due to its extensive water network in the activ e site forming secondary proton wires [4]. Carbonic anhydrase catalyzes one of the most rapid reactions; it is one of the fastest enzymes studied [1]. Its reaction speed is due, in part, by the amphiphilic nature of the active site [1]. The hydrophobic side is used to bind carbon dioxide, while the hydrophilic patch functions to optimally orient the carbon dioxide molecule for the reaction [1]. The hydrophobic wall forms a well-defined pocket near the zinc-hydroxide and is composed of the following amino acids: Val142, Val121, Leu197 and Trp208. The hydrophilic patch consists of Thr198 and Glu106, which form a hydrogen bond network with the Znà ¢Ã¢â€š ¬Ã¢â‚¬â„¢OH to stabilize and orient it for nucleophilic attack on CO2 [2]. Therefore, any modifications to the hydrophobic pocket would change its structure, and consequently, its catalytic efficiency [1]. In this study, the importance and role of His64 and Val142 to the structure and mechanism of hCAII are determined through site-mutagenesis and subsequent characterization of the new mutants, H64A (His64 Æ’Â   Ala) and V142D (Val142 Æ’Â   Asp) via kinetic and structural analysis. The changes that arise from the substitutions may prove to be applicable to drug synthesis because hCAII is known to be involved in a variety of diseases, for example, Marble brain disease, where mutations in the hCAII gene leads to a deficiency in the enzyme which is an autosomal recessive disease [5]. Studies in hCAII mutations can be used to design folding modulators to suppress misfolding which frequently occurs due to hCAII destabilization [5]. Another major disease involved with hCAII gene is osteopetrosis. The hCAII genes inactivation decreases osteoclast function in bone, and knowledge of hCAII mutations that inactivate the enzyme may lead to better understanding of bone remodelling [6]. Some carbonic anhydrase diseases use inhibitors (CAI) to suppress the hCAII as a therapeutic treatment. Inhibitors prevent hCAII activity by inhibiting either of the reaction steps: the conversion of CO2 which involves V142 in the hydrophobic pocket, or the rate limiting step, proton transfer, in which His64 is crucial. Experimental Procedure Site directed mutagenesis via the PCR-based Quick-change method was performed on hCAII as cited in Woolley (2011) for 10 ng and 20 ng wild-type plasmids (hCA2pET24b from Novagen) [7]. Table shows the sequence of the primers used in the PCR reactions. Products of PCR mutagenesis reactions were run on 0.7% agrose gels to determine size. The gels were run at 150 V in 1X TAE buffer. Red safe dye from Intron Biotechnology was used in the agrose gel instead of ethidium bromide for safety reasons [7]. The standard molecular weight ruler used was a 1 kB DNA ladder from Fermentas. Table : Primer sequences used in mutagenesis of hCAII in the forward and reverse direction for mutants H64A and V142D Mutant Direction Sequence MW (Da) %GC TM ( °C) H64A Forward GGATCCTCAACAATGGTgcTGCTTTCAACGTGGAG 10778 51 67 Reverse CTCCACGTTGAAAGCAgcACCATTGTTGAGGATCC 10709 V142D Forward CTGATGGACTGGCCGaTCTAGGTATTTTTTTG 9868 44 62 Reverse CAAAAAAATACCTAGAtCGGCCAGTCCATCAG 9779 The enzyme, DpnI, was then used to digest methylated DNA (the parent template DNA). The DNA vector that contained the mutation was transformed into supercompetent E.coli turbo cells from New England Biolabs by heat shock [7]. LB-agar plates were prepared to grow the transformed cells containing mutant genes (i.e. H64A and V142D hCAII gene) [7]. Both were injected with Kanamycin to ensure that the culture that grows will have the desired mutation [7]. A miniprep culture was set-up from the LB-agar plate into LB medium to grow one colony for DNA analysis [7]. Restriction enzyme mapping was prepared and XhoI and BglII were chosen, they were used under buffer 3 for optimal efficiency. Plasmid putification was performed using the QIAprep Spin Miniprep Kit, and then the chosen restriction enzymes were carried out and were run on 1% agrose gel [7]. A sample of the purified DNA was sent to an external company (ACGT) for commercial sequencing (Sanger dideoxy type) to verify if the mutagenesis occurred correctly. The sequence was analyzed using the program BioEdit. To determine the level of confidence of the sequencing results, the purified DNA was quantified using UV/Vis absorption via a spectrometer [7]. The concentration was calculated using Ecà ¢Ã¢â€š ¬Ã¢â‚¬â„¢1(260) as 50 ÃŽÂ ¼g/mL. Purified plasmid DNA was transformed into E.coli BL2(DE3) cells to initiate protein expression by heat shock, similar to the transformation into turbo cells [7]. The cells were cultured and a single colony was grown. Once sufficiently grown, ITPG and ZnSO4 were added to induce protein expression [7]. SDS-PAGE was used to confirm protein expression and was analyzed against an unstained protein molecular weight marker by Fermentas. The protein and ladder was stained with coomassie blue [7]. Affinity chromatography was used to purify the mutant hCAII proteins [7]. The matrix used was agrose linked to p-(aminomethyl)benzenesulfonamide, exploiting the tight binding that occurs between hCAII and sulphonamides. Once purified, the protein was dialyzed using a 6000-8000 Da dialysis membrane to replace the elution buffer with protein buffer and removes the matrix from the protein [7]. SDS-PAGE is again used to confirm the protein is still present after purification and to check its approximate molecular weight. It was run for two different amounts of protein, 2 ÃŽÂ ¼g and 10 ÃŽÂ ¼g, and also ran 10ÃŽÂ ¼L of wash fractions from affinity chromatography [7]. Protein concentration was determined by UV absorption at 280 nm in a final concentration of 6M guanidine hydrochloride. From the calculated concentrations, purity of the protein could be assessed via SDS-PAGE. To characterize this purified hCAII protein, a variety of analyses were done. Two types of mass spectrometry (MS) were performed: electrospray ionization (ESI) and matrix-assisted laser desorption ionization (MALDI) [7]. The MS analysis was used to confirm the presence of the mutation in hCAII with intact and digested protein. Protein samples (H64A and V142D hCAII) were not diluted for either of the MS analyses as cited in Woolley (2010). Samples of 10 ÃŽÂ ¼L of stock protein concentrations (37.6 ÃŽÂ ¼M H64A and 3.2 ÃŽÂ ¼M V142D hCAII) were used for analysis of the molecular weight of the intact protein by ESI-MS. Both mutants were then digested by Trypsin Gold (MS grade) from Promega and the resulting fragments were evaluated by ESI-MS as well [7]. A 50 ÃŽÂ ¼L sample was used for each mutant, 40 ÃŽÂ ¼L of the mutant at stock concentration and 10 ÃŽÂ ¼L of the Trypsin Gold. A couple ÃŽÂ ¼L of the digested mutants were saved for MALDI-MS and the rest was used for ESI-MS. Once the molecular weights for each of the digested fragments were determined by ESI-MS, the products were run through a protein database to confirm the identity of the protein and mutations [7]. The 1 ÃŽÂ ¼L of the tryptically digested mutants prepared for ESI-MS, subsequently underwent MALDI-MS. The 1 ÃŽÂ ¼L samples were mixed with a matrix consisting of 1 ÃŽÂ ¼L ÃŽÂ ±-cyano-4-hydroxycinnamic acid (CHCA) and 1 ÃŽÂ ¼L of 0.1% trifluoroacetic acid (TFA) [7]. The entire mixture was pipette onto a MALDI well and was inserted into the mass spectrometer and a MALDI-MS spectra was obtained. Michaelis-Menten kinetics was used to determine the KM and kcat of the p-nitrophenyl (PNPA) hydrolysis reaction [7]. The ionized product from the hydrolysis, p-nitrophenol (PNP) produces a bright yellow colour that was used to follow the rate of the reaction via the Perkin Elmer Lambda UV/Vis spectrophotometer [7]. Various sample concentrations of PNPA were set up to have a final enzyme concentration of 0.2 ÃŽÂ ¼M in protein buffer [7]. The initial rate measurements of each PNPA concentration were taken for wild-type enzyme, H64A mutant, V142D mutant, and a blank with no additional enzyme added (refer to data tables in Enzyme Kinetics I [7]). PNP has a molar absorption coefficient (ÃŽÂ µ) of 1.73ÃÆ'-104 Mà ¢Ã¢â€š ¬Ã¢â‚¬â„¢1cmà ¢Ã¢â€š ¬Ã¢â‚¬â„¢1. This was used to calculate Michaelis-Menten values: Vmax, KM, kcat, and kcat/KM [7]. The ability of hCAII mutants (H64A and V142D) and wild-type hCAII to catalyze the hydration of CO2 was measured. The pH of the solution was measured to track the progress of the reaction because the reaction generates protons. Enzyme solutions were prepared according to table 2 in [7]. The buffer used in the table was 50 mM TRIS buffer (pH 7.8). Additional enzyme samples were prepared for 25 nM of wild-type hCAII and 100 nM of H64A mutant in a final concentration of 22.5 and 29.92 mM imidazole buffer (pH 7.8) respectively to determine chemical rescue of mutant H64A. The pH of the CO2 hydration assay was measured using a pH probe and pH meter at 5 second increments for a total of 90 seconds starting at the beginning of the reaction [7]. The slope of the initial changes in the first 2 points was considered to be the V0 for each enzyme concentration. From the initial velocity, a kcat value can be calculated for each enzyme using the assumption that [S] >> KM, the Michaelis-Menten equati on simplifies to kcat=V0/[E]. The third kinetics experiment used fluorescence to determine the binding constant of dansyl amide (DNSA) and acetazolamide (AZ) (from Sigma-Aldrich) to H64A and wild-type hCAII was performed using the Perkin Elmer Fluorometer [7]. Stocks of 1 mM and 200 ÃŽÂ ¼M of DNSA were prepared from a 21.6 mM DNSA stock by dilution with DMSO. Enzyme stocks were diluted to 0.25 ÃŽÂ ¼M with TRIS buffer to make a 10 mL solution. A 1 mL sample of H64A from stock made was titrated with DNSA in small increments [7]. The fluorometer emissions were taken at 470 nm. AZ titration in competition with DNSA was not able to be completed. The last characterization experiment done was molecularly modelling the hCAII wild-type enzyme, as well as the mutants H64A and V142D. The molecular model of hCAII analyzed was derived by x-ray crystallography and found in the Protein Data Bank (PDB) repository. The wild-type and H64A hCAII structures examined had a PDB code of 1CA2 and 1MOO respectively. At present, no crystal structure has been found for V142D hCAII. The Swiss PDB Viewer program was used to visualize the protein structures. Secondary structures of the proteins were able to be observed. Residues around the metal active site and the Ramachandran plot were explored. Homology between hCAII and other carbonic anhydrase isozymes, hCAIV (PDB code 1ZNC) and hCAI (PBD code H1CB), were also studied by performing an iterative magic fit on the ÃŽÂ ±-carbons and structure alignment for each pair. The root mean square (RMS) between hCAII and the other isozymes were also analyzed to determine conserved and deviated regions in the structures. The binding of cobalt in the hCAII active site was also investigated (PDB code 3KOI). The structural inhibition of hCAII by AZ was also gleaned by structural analysis (PDB code 3HS4). Its mode of inhibition and binding sites were shown through the crystal structure. Lastly, the Swiss PDB Viewer program was used as a tool to theoretically synthesize mutations and compare it to the actual structure as determined by other scientists, for example, by aligning the virtual and crystallized mutations to determine deviations in structure by performing RMS. Results Site-directed mutagenesis PCR. Products from the PCR mutagenesis reactions were examined using 0.7% agrose gel electrophoresis. Two samples of differing amounts of template DNA (10 ng and 20 ng) were used for each mutant (Figure ). Bands were only observed for samples containing 20 ng of the hCA2pET24b DNA template plasmid (Figure ). The size of the bands observed coincides with the size of the plasmid used, 6018 bp. Heat shock transformation and isolation of plasmid. Several colonies were observed after plasmid transformation for both mutants, and 1 colony from each mutant was chosen for restriction enzyme digest with BglII and XhoI. Figure : Electrophoretic run on 0.7% agrose gel of DNA of hCAII mutants from PCR mutagenesis reactions. Lane 1 is the GeneRuler ladder by Fermentas and lanes 10-13 are the following: V142D (10 ng), V142D (20 ng), H64A (10 ng), and H64A (20 ng). As suggested from the gel, the mutants in the 20 ng plasmid was more successful than the 10 ng plasmids in determining relative molecular weights. Both mutants in the 20 ng plasmid show a band at approximately the 6000 base pair mark, which coincides with the number of base pairs in the hCA2pET24b plasmid that was used (6018 base pairs). Quantification of pure plasmid DNA. A 1/20th dilution was carried out on the purified DNA with elution buffer (EB; 0.1 M Tris, 0.4 M KSCN, pH 7). The absorption of the diluted DNA at 260 nm and 280 nm was taken by a UV/Vis spectrophotometer and the relative DNA purity was determined (Table ). The assumption that Ecà ¢Ã¢â€š ¬Ã¢â‚¬â„¢1 260 = 50 ÃŽÂ ¼g/mL for DNA was applied in the calculation of concentrated and diluted concentrations of purified DNA (Table ). Table : Relative DNA purity for mutants V142D and H64A determined by UV/Vis spectrophotometer absorbance at 260 and 280 nm. Calculated concentrations of mutants from absorbance data, where Ecà ¢Ã¢â€š ¬Ã¢â‚¬â„¢1 260 = 50 ÃŽÂ ¼g/mL. Mutant Wavelength, ÃŽÂ » Absorbance Units Relative DNA Purity (A260/A280) Concentrated (ÃŽÂ ¼g/mL) Diluted (ÃŽÂ ¼g/mL) V142D 260 nm 0.3117 1.7852 311.70 15.59 280 nm 0.1746 H64A 260 nm 0.2653 1.7581 265.30 13.27 280 nm 0.1509 Enzyme restriction digest. Purified plasmid DNA of mutants were digested with XhoI and BglII, separately and together in a single and double digest for both mutants. The digested and undigested samples were run on 1% agrose gel, and 2 bands were observed around the 6000 and 7000 bp marker for all 8 samples (Figure , Figure ). The expected length of the bands in the double digest should be 892 bp and 5126 bp (Figure ). Figure : Electrophoresis performed in 1% agrose gel of digested V142D hCAII in lane 1-4. The (1 kB) GeneRuler DNA ladder is shown in lane 5. Lane 1-4 contain the following: V142D plasmid, V142D + XhoI, V142D + BglII + XhoI, and V142D BglII. Double bands are shown at the 6000 and 7000 bp marker for all 4 V142D samples. Figure : Electrophoresis performed in 1% agrose gel of digested H64A hCAII in lane 1-4. The (1 kB) GeneRuler DNA ladder is shown in lane 5. Lane 1-4 contain the following: H64A plasmid, H64A + XhoI, H64A + BglII + XhoI, and H64A BglII. Double bands are shown at the 6000 and75000 bp marker for all 4 H64A samples. Figure : Restriction enzyme cut sites and position of hCAII gene (5072-5854) on the hCAI2pET24b plasmid DNA Sequencing. The mutations for both V142D and H64A in the hCAII gene were successful according to the sequenced DNA result obtained from ACGT. Other mutations in the DNA sequence were observed in both mutants, but since the aligned protein sequence was the same, mutations were likely to be silent mutations due to amino acid redundancies. When sequenced in the forward direction by T7 polymerase, a protein mutation was found (K153N) other than the desired mutation of V142D; however, when sequenced in the reverse direction by T7 polymerase terminator (T7TER), K153N was not observed. Plasmid DNA transformation into E.Coli BL21(DE3) cells. Following transformation into BL21(DE3) cells, colonies were observed for both hCAII mutants (V142D and H64A). A random colony was chosen to be cultured and then was induced to express protein with 270 ÃŽÂ ¼M IPTG and 0.1 mM ZnSO4. SDS-PAGE for protein expression. Protein expression was tested with SDS-PAGE. The expected molecular weight of V142D hCAII is approximately 29.2 kDa and the expected molecular weight of H64A hCAII is approximately 29.1 kDa. SDS-PAGE bands are observed between the ladder markers 25.0 kDa and 35.0 kDa for both mutant proteins (Figure , Figure ). Figure : SDS-PAGE loaded with V142D hCAII proteins to examine protein expression. Samples were loaded in different volumes of protein to ensure gel visualization. Lane 15 contains the Fermentas protein molecular ladder and lane 1-4 contain the following: 1 ÃŽÂ ¼L à ¢Ã¢â€š ¬Ã¢â‚¬â„¢IPTG, 4 ÃŽÂ ¼L à ¢Ã¢â€š ¬Ã¢â‚¬â„¢IPTG, 1 ÃŽÂ ¼L+IPTG, 4 ÃŽÂ ¼L +IPTG. All 4 samples had some form of protein expression between 25.0 to 35.0 kDa. Figure : SDS-PAGE loaded with H64A hCAII protein to examine protein expression. One sample was loaded with 4 ÃŽÂ ¼L of H64A protein and +IPTG in lane 10. Lane 6 contains the Fermentas protein molecular ladder. The one H64A sample loaded showed an expression between 25.0 and 35.0 kDa. Calculation of pure protein concentration and extinction coefficient. Following affinity purification and dialysis, pure protein concentration was calculated from UV absorption measurements at 280 nm and the known extinction coefficient of hCAII as 50070 Mà ¢Ã¢â€š ¬Ã¢â‚¬â„¢1cmà ¢Ã¢â€š ¬Ã¢â‚¬â„¢1 (Table ). The final concentration of the samples of V142D and H64A hCAII were 3.2 ÃŽÂ ¼M and 37.6 ÃŽÂ ¼M respectively. Table : UV absorption measurements at 280 nm of purified protein and the resulting final concentration Mutant Average A280 Protein concentration (ÃŽÂ ¼M) V142D 0.01583 3.2 H64A 0.1884 37.6 SDS-PAGE to assay purity and check approximate molecular weight. Several samples were loaded into the SDS-PAGE for each mutant protein: lysate and wash fractions (collected from affinity chromatography), 2 ÃŽÂ ¼g protein, and 10 ÃŽÂ ¼g protein. For H64A, a visible band was only observed for the 10 ÃŽÂ ¼g sample (Figure ). The band was located between the 35 kDa and 25 kDa markers on the ladder. For V142D, none of the 4 samples resulted in a band on the gel (Figure ). Figure : SDS-PAGE shown for H64A mutant protein. Lane 1 contains the Fermentas protein molecular weight marker. Lane 11-14 contains H64A samples of the following (in order): lysate, wash fraction, 2 ÃŽÂ ¼g protein, and 10 ÃŽÂ ¼g protein. Only the 10 ÃŽÂ ¼g protein had (faint) observable bands located between the 25 and 35 kDa markers. Figure : SDS-PAGE shown for V142D mutant protein. Lane 4 contains the Fermentas protein molecular weight marker. Lane 12-15 contains V142D samples of the following (in order): lysate, wash fraction, 2 ÃŽÂ ¼g protein, and 10 ÃŽÂ ¼g protein. No observable bands are seen for any of the samples. Mass spectrometry. ESI-MS was not successful in analyzing the molecular weight of intact and digested protein of both mutants. A MALDI spectrum was able to be generated for the digested proteins; however, without the digested ESI spectrum to compare to, the peaks from the MALDI spectrum can only be speculatively assigned. Kinetics: Hydrolysis of PNPA. Using the molar absorption coefficient of PNP (1.73ÃÆ'-104 Mà ¢Ã¢â€š ¬Ã¢â‚¬â„¢1cmà ¢Ã¢â€š ¬Ã¢â‚¬â„¢1), the rate of each reaction was determined. The predicted rate was calculated using the Michaelis-Menten kinetics: . The plot of predicted rates and actual initial rates vs. PNPA concentration can be seen in Figure , Figure , Figure for wild-type, H64A, and V142D hCAII respectively. The Vmax and KM values for each enzyme were calculated by minimizing the square difference between the predicted and actual reaction rates, and the kcat was calculated using the equation: (Table ). Table : Calculated Michaelis-Menten parameters for wild-type, H64A, and V142D hCAII catalyzing the hydrolysis of PNPA. Wild-type hCAII H64A hCAII V142D hCAII Vmax (ÃŽÂ ¼M/sec) 1.202 0.812 0.218 KM (mM) 1.280 1.957 8.362 kcat (sà ¢Ã¢â€š ¬Ã¢â‚¬â„¢1) 5.141 ÃÆ'- 10à ¢Ã¢â€š ¬Ã¢â‚¬â„¢3 2.159 ÃÆ'- 10à ¢Ã¢â€š ¬Ã¢â‚¬â„¢2 6.825 ÃÆ'- 10à ¢Ã¢â€š ¬Ã¢â‚¬â„¢2 kcat/KM (Mà ¢Ã¢â€š ¬Ã¢â‚¬â„¢1sà ¢Ã¢â€š ¬Ã¢â‚¬â„¢1) 4.02 11.032 8.162 Figure : Michaelis-Menten plot of initial rate vs. concentration of PNPA added for wild-type hCAII enzyme. Figure : Michaelis-Menten plot of initial rate vs. concentration of PNPA added for H64A hCAII enzyme. Figure : Michaelis-Menten plot of initial rate vs. concentration of PNPA added for V142D hCAII enzyme. Kinetics: CO2 hydration. Initial velocity (V0) values were calculated by measuring the progression of the reaction (via concentration of protons) with time (Table , Table , and Table ). kcat values were then calculated using the same equation as in the hydration of PNPA and averaged for the individual enzymes (wildtype, H64A, and V142D hCAII) in a particular buffer (i.e. TRIS or imidazole). Table : Initial velocity (V0) and kcat values calculated for the hydration of CO2 by wild-type hCAII in TRIS buffer and imidazole buffer. Wild-type concentration (nM) V0 for WT+TRIS (M/s) V0 for WT+Imidazole (M/s) 0 1.3E-08 6.05778E-08 1.5 1.1E-08 N/A 2.5 1.1E-08 5.63E-08 5 2.1E-08 5.16E-08 12.5 5.9E-08 5.63E-08 Average kcat (sà ¢Ã¢â€š ¬Ã¢â‚¬â„¢1) 5.3 ±1.62 12.44 ±9.19 Table : Initial velocity (V0) and kcat values calculated for the hydration of CO2 by H64A hCAII in TRIS buffer and imidazole buffer. H64A concentration (nM) V0 for H64A+TRIS (M/s) V0 for H64A+Imidazole (M/s) 12.5 1.4E-08 6.57E-08 25 1.4E-08 5.8E-08 50 1.7E-08 7.53E-08 Average kcat (sà ¢Ã¢â€š ¬Ã¢â‚¬â„¢1) 0.67 ±0.39 3.03 ±1.97 Table : Initial velocity (V0) and kcat values calculated for the hydration of CO2 by V142D hCAII in TRIS buffer. V142D concentration (nM) V0 for V142D+TRIS (M/s) 12.5 6.2E-09 25 5.4E-09 50 5.5E-09 Average kcat (sà ¢Ã¢â€š ¬Ã¢â‚¬â„¢1) 0.27 ±0.19 Fluorescence detection of ligand binding. DNSA was titrated with H64A hCAII to determine its affinity for the enzyme. The dissociation constant, KD, for DNSA was determined to be 0.086 ÃŽÂ ¼M when protein concentration was 0.25 ÃŽÂ ¼M. Competitive titration of H64A-DNSA hCAII with AZ was attempted, but was not successful as DNSA binding was too tight, making it difficult to be displaced by AZ. Molecular modeling. Literature models of wild-type (PDB code 1CA2) and H64A (PDB code 1MOO) hCAII were analyzed. There is no available structure of V142D hCAII at present. The secondary structure of wild-type is composed of 18 ÃŽÂ ²-sheets (77 residues) and 10 ÃŽÂ ±-helices (42 residues), with the majority of the ÃŽÂ ±-helices falling in the domain of right-handed helices, while very few show left-handed helical properties according to the Ramachadran plot. It also seems that the active site is solely composed of ÃŽÂ ²-sheets, and no ÃŽÂ ±-helices (Figure ). Analyzing PDB structure 3HS4 (AZ bound hCAII), the mechanism as to how AZ inhibits hCAII function can be seen. AZ has 3 binding sites, 2 are novel binding sites and the other provides a mechanism of inhibition. AZ binds the zinc directly at the active site, displacing crucial ligands needed for catalysis. There were some discrepancies found between the crystal structure of H64A [1MOO] as cited on PDB and virtually mutated H64A from wild-type hCAII, resulting in a RMSD (root mean square deviation) of 0.29 Ã… (Figure ). Since no literature structure of V142D is available, no comparison between virtual and crystal structures could be made. Figure : Secondary structure of wild-type hCAII overlain with ribbon to visualize the higher arrangement. Figure : RMSD between H64A hCAII virtually mutated and literature crystal structure. Blues denote the same or similar residues, while reds and oranges indicate completely different amino acids. Discussion Agrose gel results were only visible for samples that contained 20 ng of the plasmid template DNA, rather than the 10 ng plasmid. This may be a result of more amplification during PCR with the 20 ng plasmid, and so would intensely be more visible. Though the 20 ng samples showed bands at the appropriate 6000 bp mark, there was also a faint band that can be seen near the end of the gel. This may be due to non-specific primer annealing. Quantification of DNA purity was done by exploiting the peak absorbances of protein and DNA. DNA maximally absorbs at 260 nm, while protein dominantly absorbs at 280 nm. The purity ratio reports the relative amount of DNA compared to protein present in the sample. The purity of both mutants were approximately 1.8, which is regarded as a relatively pure sample; however, a purity ratio of more than 2.0 would have been ideal. The restriction enzyme digest showed 2 bands (7000, 6000 bp) for all samples, which may have been a sign of poor mixing/ pipetting since the volumes of restriction enzyme were extremely small amounts. If this is the case, only some of the DNA was nicked and some were not, which would result in 2 bands. It was expected that the plasmid sample would have a high band (supercoiled), each of the singly digested samples would have a slightly lower band (nicked), and the doubly digested would show 2 bands that indicated the fragment size of 892 and 5126 bp. Sequencing results showed that a protein mutation occurred when the sample was sequenced in the forward direction by the T7 polymerase. A lysine at position 153 had mutated to glutamine (K153N). However, this mutation was not observed when the T7 polymerase terminator was used to sequence the sample in the reverse direction. A mutation that occurs in one sequencing direction and not the other is usually attributed to sequencing errors, which may be the reason in this case. The SDS-PAGE bands for protein expression coincided with the expected molecular weight for both mutants, which could suggest that the correct proteins were expressed; however, there is a possibility that the proteins expressed could be of similar weight, but completely different. Interestingly, the V142D samples that did not include the protein inducer, IPTG, had a more intense band than the faint ones found for the samples that did include IPTG. This may just be a result of mislabelling. The SDS-PAGE performed to assess purity after the purification process. Mutant V142D had low protein expression as evidenced by its concentration of 3.2 ÃŽÂ ¼M. The V142D mutant should have very low protein expression according to Fierke et al. (1991) because valine at position 142 is uniquely required for maximal expression in E.Coli. It is suggested that by altering position 142, protein stability decreases [2]. Therefore, the protein that was expressed in the previous SDS-PAGE gel may not be V142D hCAII at all. The sample may have been small fragmented contaminant proteins that would have completely run off the gel altogether. However, the low concentration of V142D after purification may also be a major factor in the lack of gel bands observed as well. Unlike V142D, H64A hCAII concentration should not have affected its lack of bands because it was calculated to have had a reasonable concentration of 37.6 ÃŽÂ ¼M. There were some problems loading the samples into the wells; t his could be an explanation as to no observable gel bands. ESI-MS is dependent on concentration because it affects the size of primary droplets [8]. The unsuccessful determination of molecular weight of V142D hCAII may be attributed to its low concentration. The H64 hCAII mutant was also not able to be successfully analyzed with ESI-MS. A possible reason for the failure was that it was not kept on ice while it was not being used. The enzyme may have become inactive and degraded into smaller fragments. This would explain the ESI-MS output obtained for H64A. No definite molecular mass was determined, but the spectrometer did detect a lot of small protein fragments in the sample, all under 1000 amu. The kinetic values obtained from PNPA hydrolysis do not follow similar trends found in literature [2]. The kcat/KM for wild-type hCAII (2500 ±200 Mà ¢Ã¢â€š ¬Ã¢â‚¬â„¢1sà ¢Ã¢â€š ¬Ã¢â‚¬â„¢1) was found to be significantly larger than V142D hCAII (3 ±0.3 Mà ¢Ã¢â€š ¬Ã¢â‚¬â„¢1sà ¢Ã¢â€š ¬Ã¢â‚¬â„¢1) in literature, more than 800ÃÆ'- larger [2]. Experimental calculations yielded kcat/KM for V142D (8.16 Mà ¢Ã¢â€š ¬Ã¢â‚¬â„¢1sà ¢Ã¢â€š ¬Ã¢â‚¬â„¢1) to be about 2ÃÆ'- larger than wild-type (4.02 Mà ¢Ã¢â€š ¬Ã¢â‚¬â„¢1sà ¢Ã¢â€š ¬Ã¢â‚¬â„¢1), which did not follow literature patterns. The literature trends make more biolog

Creative Commons Essay -- Music Musical File Sharing Essays

Creative Commons Creative Commons encourages artists to share and distribute their work for free. And that could be the key to a new multibillion-dollar industry. People can widely redistribute other people works, as long as they provide the credit to the authors; create new works based on an existing ones, provided they offer those derivations back to the public on the same terms. This paper analyzes the conflicts between the need of technology for creativity and innovation versus the legal aspect of copyright. An alternative emerging approach for licensing music, software, research paper and many other resources on the internet, creative common sharing copyright, is introduced. The paper does not analyze if creative commons is ethical or not; the answer will be yes under all ethical approaches. An ethical analysis on intellectual property, using multiple approaches, is instead presented. Technology and Legal Trend Restriction imposed by intellectual property law, for someone like Laurence Lessig, chairman is a professor of law at Stanford and founder of the School's Center for Internet and Society, have run out of control. The restriction impose by the market and by the state are affecting and threatening internet. Changes in the copyright and patent laws and the regulation of broadcast seem to protect the interest of few against new way of doing things. Law and technology are constantly increasing control on the uses of creative works at a level never seen in history before. The Digital Millennium Copyright Act (DMCA) does not permit, under the law, the usage of copyrighted material for fair use. Bill Gates recently stated that "If people had understood how patents would be granted when... ...andy Starr, The Creative Commons, February 2004, http://www.spiked-online.com/Articles/0000000CA401.htm 4. Joe Kaplinsky, Who owns ideas?, Frebruary 2002, http://www.spiked-online.com/articles/00000002D42C.htm 5. Sandy Starr, Copycat copyright, March 2003, http://www.spiked-online.com/articles/00000006DCC2.htm 6.Chris Evans, Tightening the net, June 2002, http://www.spiked-online.com/articles/00000006D929.htm 7.Dwight Duego, The boundary of Intellectual Property, January 2004, http://www.adtmag.com/article.asp?id=8778 8. Mark Alfino, Intellectual Property and Copyright Ethics, Business and Professional Ethics Journal, Vol.10 No.85, http://guweb2.gonzaga.edu/faculty/alfino/dossier/Papers/COPYRIGH.htm Useful Links 1. www.creativecommons.org 2. http://www.lessig.org 3. http://cyberlaw.stanford.edu/ 4. http://cyber.law.harvard.edu/home/

Poverty in Ghana

In recent years, Ghana has emerged as a leading country in the Western and Central Africa region. It has developed its economy on a scale that could enable it to meet the Millennium Development Goals (MDGs) before the 2015 deadline. It also managed a smooth and peaceful political transition in 2008 and 2009, and has created a political and policy environment conducive to economic and social progress and poverty reduction. The Ghanaian economy has grown at an average annual rate of 4. 8 per cent over the past two decades. By 2008 GDP growth had reached 7. per cent. The agriculture sector, which contributed 33. 5 per cent of GDP in 2008, remains the country’s major engine of economic growth. Rapid economic progress has all but halved national poverty rates, which have fallen from approximately 50 per cent in 1991 to 28. 5 per cent in 2006. In the last decade, poverty rates dropped by 8. 6 per cent in urban areas and by 10. 4 per cent in rural ones. Ghana’s growth and poverty reduction rates are probably the best that have been achieved throughout sub-Saharan Africa in the past 15 years. Where are Ghana's rural poor people? Although there has been a substantial overall decline in the incidence of poverty in Ghana, poverty still has a firm grip on rural areas, especially in the north. There is a wide disparity in income between people living in the drought-prone northern plains, and those living in the south, where there are two growing seasons and greater economic opportunities. Who are Ghana's rural poor people? Just over half the country’s population lives in rural areas. The poorest parts of Ghana are the savannah regions of the north (the Northern, Upper East and Upper West regions), where chronic food insecurity is widespread and livelihoods are more vulnerable. Poor rural people have limited access to basic social services, safe water, roads that are accessible year round, and electricity and telephone services. Poverty is most severe among food crop farmers, who are mainly traditional small-scale producers. About six in ten small-scale farmers are poor, and many are women. Women bear heavy workloads. In addition to their domestic chores, they are responsible for about 60 per cent of agricultural production. More than half the women who head households in rural areas are among the poorest 20 per cent of the population. Why are Ghana's rural people poor? According to the Government’s poverty reduction strategy paper, low productivity and poorly functioning markets for agricultural outputs are among the main causes of rural poverty. Small-scale farmers lack the technologies and inputs, such as fertilizer and improved seed, that would increase yields. Population pressure leads to shorter fallow periods or even continuous cultivation in the densely inhabited Upper East and Upper West regions, causing soil erosion and loss of fertility. Land degradation poses a long-term threat to farmers’ livelihoods and incomes. Only a small proportion of farmers have access to irrigation. Land ownership and land security are regulated by complex systems that vary widely. Many farmers lack rural infrastructure and equipment for storing, processing and marketing their products. In order to overcome their vulnerability and food insecurity, poor rural populations need help in sustainably increasing their incomes. Good opportunities exist to link farmers to markets and to modernize agriculture. The rural private sector could play an important role in making farming a profitable business through access to financial services, farm inputs and linkages to agroprocessors and traders. www. cfr. org Ghana is rich in natural resources and has one of the strongest emerging economies in Africa. The capital city, Accra, is one of the wealthiest and most modern cities on the continent, and is experiencing a period of rapid growth. Ghana is rapidly urbanizing. Despite this, most of Ghana's poor live in rural areas without basic services such as health care and clean water. Small-scale farmers, who are affected most by rural poverty in Ghana, depend on outdated farming tools and lack access to improved seeds and fertilizers to increase crop yields. Our Work The Hunger Project has been working in Ghana since 1995 and is empowering partners in 49 epicenter communities to end their own hunger and poverty. Through its integrated approach to rural development, the Epicenter Strategy, The Hunger Project is working with partners to successfully access the basic services needed to achieve the Millennium Development Goals (MDGs) and lead lives of self-reliance. Scaling-Up the Epicenter Strategy Ghana is the first country in which The Hunger Project is scaling up its Epicenter Strategy to provide coverage to a set geographical area and reach significantly more people. In July 2006, The Hunger Project received a US$5 million investment to scale-up Hunger Project work in the Eastern Region of Ghana. By the end of the five-year commitment, The Hunger Project-Ghana aims to have mobilized enough epicenter communities so that each villager in the Eastern Region is within walking distance to an epicenter building. By the end of the second year of its scale-up program, The Hunger Project-Ghana had successfully completed construction of 12 epicenters. Scaling-up has presented a number of challenges to The Hunger Project-Ghana, such as acquisition of land and the mobilization of construction materials. Our staff, however, is equipped with the training and skills to continue to mobilize partners to end their own hunger in the Eastern Region. Empowering Women The Hunger Project-Ghana has pioneered the Women's Empowerment Program (WEP), which empowers women to become strong leaders in their households and communities. The WEP is a series of workshops that focus on legal, civic and reproductive health rights as well as leadership skills for selected women. The trained women, also known as â€Å"animators,† then carry out community-based educational activities using drama, mini-lectures and discussions. These animators are also trained to provide counseling and distribute non-prescriptive contraceptives. Microfinance Through its Microfinance Program in 2009, The Hunger Project disbursed 1,834 loans totaling $146,421. Partners in Ghana deposited $23,589 in savings during the year. Of the epicenters that are operating in Ghana, three have government-recognized Rural Banks and are self-reliant.

A Person Who Has Made a Great Impact on My Life

Sometimes we come across a person who changes our whole perspective of life for better. Last summer I met Uncle Joe. My parents sent me to my uncle on holidays. I was supposed to spend my whole summer holidays with him and his family. That was the first time we have been introduced to each other. At first, I thought that it was going to be my worst holiday ever. I felt embarrassed of living with people I hardly knew and wanted to return home but days passed and I got to know my uncle better. Everyday, we used to wake up early in the morning and go for fishing.We packed our bags with provision and went to the river. He taught me how to fish and then we sat together for hours, fishing. While fishing, he used to tell a lot of interesting stories based on his own life experience. He told me stories about his student life, about his career, and about life in general. Once he asked me what I want to become and what I want to achieve in my life. I did not know what to answer  and never th ought about it before. He told me that I should  decide wisely  so that in the end I would not regret about anything I have done. I  told him  that I don't know what I want to do.Then he advised that I should do only what  I am interested in. I remember his words, he said, â€Å"Don't care about money or prestige of your occupation. Just do what you really love. † I started to list down the things which I really love and began writing them in my diary. The whole summer I spent exploring myself and finally deciding that I want to connect my future with physics and architecture. Therefore, I came to conclusion that I should study civil engineering. But this is not all what my uncle has helped me with. He also told me that I should be determined and persistent in achieving my goals.He often said that resolute determination is the truest wisdom. However, I think the most important lesson I got from my uncle is to see only positive side in any occasion. One day, we did n ot catch any fish and went home with empty hands. I looked at my uncle and noticed that he was smiling. I wondered why he was smiling as I did not see any reason to be happy. Then he said exuberantly, â€Å"We didn't catch any fish, but we had a great time. I don't see any reasons to be unhappy. † I admire his attitude. He is always grateful to everything he has and is truly happy.Uncle Joe is the person who has made an impact on my life. After that holiday, I changed my lifestyle and attitude. I used to spend a lot of time in front of my computer, playing games and surfing the net. Now, I spend more time studying, playing basketball and going out with my friends. I learned a lot of things which were new to me. I became more serious, optimistic and mature. During the time I spent with him, I also learned that no matter how smart I am or how much I know, there is always more to learn or experience. I'm grateful to him that he has helped me to find out who I am and what I want.

Never Found The Time Essays - Free Essays, Term Papers

Never Found The Time Essays - Free Essays, Term Papers Never Found The Time Author unknown Submitted by Jaz ******** I knelt to pray but not for long, I had too much to do. I had to hurry and get to work For bills would soon be due. So I knelt and said a hurried prayer, And jumped up off my knees. My Christian duty was now done My soul could rest at ease. All day long I had no time To spread a word of cheer. No time to speak of Christ to friends, They'd laugh at me I'd fear. No time, no time, too much to do, That was my constant cry, No time to give to souls in need But at last the time, the time to die. I went before the Lord, I came, I stood with downcast eyes. For in his hands God held a book; It was the book of life. God looked into his book and said.. Your name I cannot find. I once was going to write it down...but never found the time.

Free Essays on The Marriage of Heaven and Hell

In The Marriage of Heaven and Hell, the famous British poet William Blake wrote that â€Å"without contraries there is no progression - Attraction and repulsion, reason and imagination, and love and hate are all necessary for human existence† (Blake 122). As Blake noted, the world is full of opposites. But, more importantly, these opposites allow the people of the world to see themselves and their thoughts more clearly. For, as Blake asserts, without attraction, one cannot understand repulsion, and without imagination, one cannot understand reason. In Much Ado About Nothing (MAAN), William Shakespeare uses this idea of the power of opposites to show the differences in two types of love. Using the relationship, language, and actions of Hero and Claudio as a foil against those of Beatrice and Bene*censored*, Shakespeare has painted a world in which the ideas of courtly love only serve to illuminate those of true love. In an essay on Chaucer’s Canterbury Tales, William Kittredge defined the idea of courtly love that is illustrated in MAAN. Kittredge said that courtly love must involve a love that is extremely idealized and superficial, with the vassal or servant-like suitor, who is often a valiant knight, devoting himself completely to an ideal woman who is often the daughter of a powerful man (Kittredge 528-529). When this definition is applied to the relationship between Hero and Claudio in MAAN, one is able to recognize a perfect match. For example, Claudio, a young lord of Florence, is a valiant soldier as is shown in the first scene of the play with the comments made by the Messenger: â€Å"[Claudio] hath borne himself beyond the promise of his age, doing in the figure of a lamb the feats of a lion† (Shakespeare 1.1.11-12). He is, from the very beginning of the play, hopelessly in love with Hero, but that love is a relatively superficial thing. This is proven by the fac t that he ea! sily believes others comments abo... Free Essays on The Marriage of Heaven and Hell Free Essays on The Marriage of Heaven and Hell In The Marriage of Heaven and Hell, the famous British poet William Blake wrote that â€Å"without contraries there is no progression - Attraction and repulsion, reason and imagination, and love and hate are all necessary for human existence† (Blake 122). As Blake noted, the world is full of opposites. But, more importantly, these opposites allow the people of the world to see themselves and their thoughts more clearly. For, as Blake asserts, without attraction, one cannot understand repulsion, and without imagination, one cannot understand reason. In Much Ado About Nothing (MAAN), William Shakespeare uses this idea of the power of opposites to show the differences in two types of love. Using the relationship, language, and actions of Hero and Claudio as a foil against those of Beatrice and Bene*censored*, Shakespeare has painted a world in which the ideas of courtly love only serve to illuminate those of true love. In an essay on Chaucer’s Canterbury Tales, William Kittredge defined the idea of courtly love that is illustrated in MAAN. Kittredge said that courtly love must involve a love that is extremely idealized and superficial, with the vassal or servant-like suitor, who is often a valiant knight, devoting himself completely to an ideal woman who is often the daughter of a powerful man (Kittredge 528-529). When this definition is applied to the relationship between Hero and Claudio in MAAN, one is able to recognize a perfect match. For example, Claudio, a young lord of Florence, is a valiant soldier as is shown in the first scene of the play with the comments made by the Messenger: â€Å"[Claudio] hath borne himself beyond the promise of his age, doing in the figure of a lamb the feats of a lion† (Shakespeare 1.1.11-12). He is, from the very beginning of the play, hopelessly in love with Hero, but that love is a relatively superficial thing. This is proven by the fac t that he ea! sily believes others comments abo...

Idioms and Expressions - Eye

Idioms and Expressions - Eye The following idioms and expressions use the noun eye. Each idiom or expression has a definition and two example sentences to help with  these common idiomatic expressions. Apple of Someones Eye The expression  apple of my eye  is often used when referring to family members, or those who are closest to us to mean that they are someones favorite person or object. Jennifer is the apple of her fathers eye. Hes so proud of her. My Mercedes is the apple of my eye. Birds-eye View Birds-eye view  refers to a viewpoint from which one can see a wide area. This idiom  is often used figuratively to mean that someone can see a situation from a wider perspective. His birds-eye view of the market will help us beat our competitors. The hotel provides a lovely birds-eye view over the bay. Catch Someones Eye Catch someones eye indicates that someone or something has  gained attention. I caught the servers eye. Hell be with us shortly. That house on Elm street certainly caught my eye. Should we talk a look inside? Cry Ones Eyes Out Crying ones eyes out  is an idiom used for very sad events in ones life. It means to  cry for a very long time in a desperate manner such as at the loss of a loved one. I think you just need to cry your eyes out to get it all out of your system. Marias crying her eyes out. I wonder what happened? Eagle Eye Someone with an  eagle eye  has  the ability to see important details and notice mistakes. Show it to the editor. She has an eagle eye and will catch any mistake. Luckily, Toms eagle eye saw the discounted sweater I was looking for. Feast Ones Eyes on Something If you feast your eyes on something,  you  enjoy the sight of something. This idiom is often used to boast about a possession of which you are very proud. Feast your eyes on my new watch. Isnt it beautiful?! I cant stop feasting my eyes on my new car. Get a Black Eye If you get a black eye, you  receive a bruise from something around the eye. This idiom can  also be used figuratively to mean to suffer a defeat. I got a black eye when I bumped into the door. I guess we got a black eye trying to compete with that big corporation. Get Stars in Ones Eyes Some young people get stars in their eyes  because they  become obsessed about show business. Ever since Janet got the lead role in the high school play, shes got stars in her eyes. Just because you are handsome doesnt mean you need to get stars in your eyes. Give Someone the Eye People will run when  you give them the eye  because you  look at someone in an accusatory or disapproving manner. The teacher was giving me the eye during the test. I guess he thought I might cheat. Dont give me the eye! Youre the one who caused this mess. Have Eyes Bigger than Your Stomach Unfortunately, its easy to put on weight if you  have eyes bigger than your stomach  because you  desire more food than you can eat. Little children tend to have eyes bigger than their stomachs. I remember one crazy night when my best friend had eyes bigger than his stomach. He ordered more than six different meals! Have Eyes in the Back of Ones Head If you have eyes in the back of your head, you are  be able to see what is going on. This can be frustrating because others might think they are being secretive and are not noticed. My mom had eyes in the back of her head. I never got away with anything. Do you have eyes in the back of your head? How did you notice that? Hit the Bulls-eye When a person hits the bulls-eye,  they  hit the center of the target. This expression is also used figuratively to express an impressive result. I think we hit the bulls-eye with our new product line. Youve hit the bulls-eye by getting that job. In the Public Eye If you are in the public eye, you are  in a position where the public can observe your actions, so be very careful! Youll be in the public eye if you take that job. Hollywood actors are all in the public eye. Keep Ones Eye on the Ball People who can  keep their eye on the ball  remain concentrated, especially in a work situation. You need to keep your eye on the ball, no matter how long it takes, to succeed. His ability to keep his eye on the ball assured his eventual success. Turn a Blind Eye to Someone or Something Unfortunately, some people turn a blind eye to someone  and show that they are  willingly to ignore  something wrong. Just turn a blind eye to Ted. Hell never change. Im going to turn a blind eye to that problem for the moment. Without Batting an Eye Most parents help their children without batting an eye  because they  do it without hesitation. He purchased the $2 million home without batting an eye. John made the decision without batting an eye. Idioms With Eye Quiz FIll in the gaps with a word to complete these sentences with expressions using  eye: Our boss has an ______ eye because he catches mistakes that others miss.Lets take a ______ view of this situation to make sure we dont miss anything.Its surprising how many young people get ______ in their eyes and move to Hollywood to start a career.I ordered this cake, but its too much. Im afraid I have eyes that are bigger than my ______.My daughter is the ______ of my eye.I think you hit the ______ when you made that investment. Today, you are a millionaire!She gave her daughter $500 without ______ an eye because she trusts her to spend it wisely.Would you please stop giving me the ______! Youre making me nervous!I got a ______ eye when I fell last week.Politicians are always in the ______ eye.   Answers eaglebirds-eyestarsstomachapplebulls-eyebattingeyeblackpublic

Consumer Behaviour in UK and China Automobile Industry Dissertation - 1

Consumer Behaviour in UK and China Automobile Industry - Dissertation Example These five questions are very important as it covers all major areas for studying consumer behaviors. In order to cater to those five questions, this paper has presented and discussed important sections like literature review, primary research and secondary research. The literature review has presented an  important theoretical background to understand the mechanism of consumer behavior. The primary research has obtained important data for identifying the prevailing consumers’ behaviors and secondary research has provided the factual data to link with the primary data. The gathered primary data has disclosed major trends in consumer behavior in car markets of the UK and on the Chinese people. These primary data are also discussed and analyzed in the light of the given theoretical background and secondary data. The output of the primary data analyses has supported the consumer behavioral theories and secondary data. As per the discussions based on literature review, primary a nd secondary data, this paper is able to cater to the five pre-defined research questions. The consumer behavior in the  automobile industry of China and UK is influenced by a number of factors like culture, economic condition, brand, quality, price etc and these factors vary as per the national culture and system of the two countries. The successful automobile companies presented in both countries are able to differentiate and identify the multiple factors influencing the consumer buying behavior. The cultural paradigm of different markets like China and Britain reflect significant impacts on the purchasing behavior of cars and domestic vehicles. The main aim of the paper revolves around understanding the reasons or factors which bring about cultural changes and thereby affect the demand for cars in the two markets.  

The Holocaust and the German People Essay Example | Topics and Well Written Essays - 2000 words

The Holocaust and the German People - Essay Example No one wants to believe or less admit that a civilized world would sit back and watch a Holocaust of such proportions and still do nothing. So many, even to this day, refuse to believe and continue to deny the facts; deny the truth and claim ignorance. The sad reality is the Holocaust did occur and sadder still is the fact that all saw what was happening and few had the courage to step forward and make any attempt to stop it or even say "This is wrong." "'The Holocaust' or 'Shoah' refers to the systematic annihilation of six million Jewish people by Germany's Nazi regime over the period January 30 1933 to May 8 1945." (Overview, undated) In examining the history of the world, there have always been instances of hate and persecution, not just against the Jewish people, but against many different groups of people. The Holocaust was, however, unique in that never before and never since had there been such a systematic attempt to anneliate an entire population of people by a sovereign nation. Although during the period the Jewish people were not the only group targeted for destruction by the Nazi regieme; the list also included "Gypsies, homosexuals, political dissidents and the intellectually and physically disabled" (Overview, undated); this period will always be remembered as the attempted decimation of the Jewish population. To fully examine what was known and understood by the German people during this period and in an attempt to understand how and why it occurred, we need to look to the past and first examine the era predating the Holocaust. Events do not occur independent of one another. Therefore, to see how this occurred, why it occurred, and why the German people and the world stood by, we first need to examine the culture of the period leading up to the Holocaust. Pre 1933 Jewish people have been living in Europe for over 2000 years. Throughout their history as a people, there had always been periods of persecution and hatred. "The place of Jews in the wider society of Europe had always been characterised as a kind of exile. Certainly Christian Europe saw the Jews in their midst as rebels against their 'true' religion, responsible for the death of Christ and generally as an evil presence." (Jewish Life, undated) There have been previous instances where the persecution of the Jewish people included being confined to ghettoes, wearing distinctive markings on their clothing and other forms of degradation commonly associated with the Holocaust; however, the scope and magnitude of that period had never come close in comparison. Prior to 1933, the Jewish people living in Germany had begun to feel increasingly that they were becoming a part of mainstream society. The post World War I era, saw a real integration of the Jewis

Relationships between learning disabilities and the impact of Dissertation

Relationships between learning disabilities and the impact of behavioral disorders on standard experiences - Dissertation Example Is it possible to make an economic case for the value of government intervention in this regard? There is considerable evidence that a lesser investment will reliably produce lesser results in terms of education. (Inman, 1986). The time to start educational assistance for those with special needs is in their youth. A variety of studies indicate a principle of diminishing returns for efforts made to educate any useful job skills fully formed adults both academically and for a specific vocation. (Furstenberg, 1987) those with physical disabilities may still prove to be intellectually quite proficient. They may even rely more upon intellect than physical activity, and may be able to develop their intellectual acumen even further than the general population. If given special needs support in early childhood. Even those with mental disabilities may still grow into productive citizens if the nonfunctional can be taught to function by trained, competent specialists committed to the outcome of special education. DISCUSSION In terms of educational policy; a vital research question becomes: Should Learning Disabled and Behavioral challenged students be excluded from classes with baseline students? The challenges for these children are numerous, there are many reasons why children experiencing learning disabilities might fail in school, far beyond merely the obvious. When a child perceives him or herself as being mentally deficient/learning disabled the potential for great insecurity develops. School in general may seem painful, an embarrassing waste of time and venue for potential bullying and the degradation of the students self-worth. It is a daunting obstacle; how to focus on learning, if you were such a student – labeled and ridiculed for what it seems you have no control over. Often times, measures used in the past to give these students focused attention; placing them in segregated units, or separate institutions all together unintentionally exacerbated the problem. It makes sense to give extra attention to the learning disabled; and one might logically conclude that this attention should take place within a place all their own. But this natural exclusionary tendency towards those with behavioral, or intellectual disabilities can become a disservice, as will be demonstrated later. The findings of several researchers demonstrate how behavioral problems will disrupt the already unfavorable prospects of the learning disabled. It is worthwhile to specify the definitions of learning disabilities. Although a precision diagnosis and prognosis may not be consistent, there are warning signs that signify disability: Difficulty in learning the alphabet Poor reading comprehension Frequent mistakes when reading aloud Spelling difficulty Inability to express thoughts by writing Late language acquisition Lack of comprehension of humor, or satire Mispronunciation Confusion with written numbers Confusion with the sequence of a story (Medinenet.com, 201 1) In essence, the typical tasks that are begun upon entrance to the public school system are those the learning disabled are most challenged by. A true disability must be diagnosed with care; in past generations, so-called intelligence tests might simply measure cultural references, biasing them against other ethnicities and cultures. (Dove, 1971) An

The Marketing Strategies and Competitive Advantage Assignment

The Marketing Strategies and Competitive Advantage - Assignment Example The inter-brand differentials while volume-based sales (10 pounds of Product each week) are given 10% of retail price while below it have 7% share making the intra brand competitions more acute.   The marketing strategies are formulated by a business to achieve its business objectives in a specific market. Thackeray et al. (2008) suggest that the marketing strategies are designed to increase sales, and achieve a sustainable competitive advantage for itself. The marketing strategies of Candy Flake and its extension of Chocofill are existent in the north east markets of US with a growing potential in southern Canadian markets. Therefore the marketing strategies for the business are developed in accordance with these markets where the business also keeps the scope of e-commerce sales in the international market open. Talke & Hultink (2010) suggests that the tasks and achievements of a strategy are based out of measured outcomes over a multiyear period with specific actions and tactical plans. The brands mission and vision statements are kept in mind while designing the plan. The objective of the business is to operate and fulfil the visions that have been set forth for itself. Further, Hallback & Gabrielsson (2013) argues that the marketing strategies are based on market dominance plans of the brand, where each phase of business includes different elements in its strategy formulation. The product of Candy Flake is been launched in the market and is in a primary phase of its business life cycle. Therefore the launching strategies would include the branding, pricing and distribution plans. The brand is the name of a product offered which implies a certain values and offerings to the market (Mishra, 2012). The mission of Candy Flake suggests the business wants to project as the major stakeholder in the US and Canada market and reap the benefits from the candy industry for next 5 years. The launching of the product have the objectives of creating an identity for itself.

Paediatric Trauma Essay Example | Topics and Well Written Essays - 2000 words

Paediatric Trauma - Essay Example Details of the case are presented in this paper as well as the proper medical management of this case base on the paramedical standard of paediatric trauma protocol and other reliable sources, utilising theories and related literatures significant in this study to convey a realistic and valid approach of management. Description of the Scene At 08:49, an accident was reported involving a 9-year-old boy, named Martin, who has been hit by a car travelling approximately 30 kilometres an hour whilst crossing the road on his bicycle. On the scene, the boy is conscious with some respiratory distress 10 minutes after the incident. His left leg appears to be deformed, more likely associated to fractured tibia, fibula, or both. A man holding the boy’s helmeted head, maintaining spinal alignment and who claimed that he is the driver of the car and a trained first aider, said that Martin complained about tightness in his chest and that he has asthma. Paediatric Trauma To start with, it is important to emphasize that the term â€Å"trauma† is being used in this study according to its medical definition as a serious or critical bodily injury, wound or shock (MedicineNet.com, 2011). Trauma is the leading cause of childhood mortality because after the age of 5 years, rapid neuromuscular development, frequent social interactions with other children, wider range of activities, and less parental supervision predisposes them to greater risk. Paediatric trauma is more difficult to handle than in adults as presented in Staheli’s (2003, pp. 105-107) ‘Pediatric Orthopaedic Secrets’ book of the contributions made by Dr. Cummings about child versus adult trauma management: Details about the child’s injury will be difficult to gather if the trauma was not witnessed by an adult; assessment is difficult to establish as children frequently resist examination. Trauma causes more death and injury in children than in adults and unintentional injury caus ing blunt trauma is more common to children. Between 30% to 70% of trauma deaths in children are due to head injury because anatomically, children’s head is larger in proportion to the body than in adults and their neck muscles are weaker, thus the cervical spine is more susceptible to injury; their skulls are thinner, and scalps are more vascular than those of adults. Children’s liver and spleen are larger relative to their size than in adults, making them prone to hemorrhage due to intra-abdominal injury. Children are more prone to hypothermia due to higher body surface-to-mass ratio, their skin is thinner and they have smaller stored fats compared to adults. Children cannot be relied on when it comes to immobilization required in fracture healing during recovery as they tend to resume normal activity (if unattended) once they do not feel pain anymore. Assessment of Paediatric Trauma At the moment the health care provider reaches the scene, it is a standard operating procedure that he/she will introduce himself/herself in the crowd while conducting the primary survey and clearing the site for potential harm. Assessment of paediatric trauma in pre-hospital setting should be done rapidly to make sure that the injured child will be transported right away to the nearest facility where in-depth assessment, management, and treatment shall be given. In the pre-hospital setting, the rapid assessment and preparation includes: (1) minimizing scene time, (2) controlling external bleeding and shock, (3) immobilization, (4) immediate transport to the closest appropriate facility, (5) obtaining information from the handover, (6) assessment for level of consciousness, respiratory rate, systolic blood pressure, revised trauma score, and paediatric trauma score, (7)

Nursing Education System Assignment Example | Topics and Well Written Essays - 750 words

Nursing Education System - Assignment Example According to Chen (1996, pp. 129-149), the first collegiate nursing program was started by Peking Union Medical College (PUMC). This was a five-year program established in 1920. The first secondary level nursing school was established in 1930. The PUMC program remained the only tertiary program in Chinese education until 1952 when the communist government restructured the education system to accommodate the higher education system. However, the political upheavals delayed the implementation of the higher education program until 1980s (Xu et al., 2000, p. 208). Nursing education in Poland was established later than in China, i.e. 1911 (Sztembis, 2006, p. 102). However, the development rate of nursing education in Poland was faster compared to that of China. Polish nursing education ceased in 1914 with the rise of the First World War (Sztembis, 2006, p. 102). According to Wolska-Lipiec (1987, p. 42), most of the teachers and graduates of the school joined the military to train the soci ety on the modalities of caring for the wounded and participating in the civil sanitation. Secondary education in nursing was introduced in 1960s, while the university-based programs were introduced in 1980s (Sztembis, 2006, p. 104). Government and Nursing Organizations Influencing Nursing Education: Comparison of Poland and China. Nursing education in China was predominantly influenced by the missionaries (Xu et al, 2000, p. 207). The horizon for the Chinese health care has been expanded by programs such as the WHO collaborating center in Nursing and Midwifery, Heart to Heart and the Project Hope which have been sponsored by the government and other non-governmental organizations (Xu et al, 2000, p. 208). The prominent source of financial support is China’s Medical Board (CMB), which is an offshoot of Rockefeller Foundation (Xu et al., 2000, p. 212). CMB provides resources for faculty training, purchasing the reading materials and capacity building. In Poland, the programs a imed at preparing women for care of patients were funded by the collaboration of the American Red Cross and Rockefeller Foundation (Sztembis, 2006, p. 103). The Polish Nursing Association undertook the process of redesigning nursing through seeking support from the politicians and leaders. The 1990s witnessed the establishment of the Independent Nursing Section in the Ministry of Health which later culminated into the Department of Nursing. The self-governing body of Polish nurses, i.e. Nursing Chambers, acts as the formal representation of the nurses at the national level. Current System of Nursing Education: Comparison of Poland and China The Chinese system requires a candidate to go through the secondary nursing programs; Zhuake programs, which are equivalent to the associate degree programs in the U.S., and Baccalaureate programs for them to be registered nurses (Xu et al., 2000, p. 209). The secondary nursing program acts as a basic academic unit of a nursing school. The second ary nursing programs occur in two forms; the first, which enrolls high school graduates and a second that admits the middle and junior high school graduates. Post secondary nursing program in China has three levels: Baccalaureate, Zhuanke and Graduate (Xu et al., 2000,

Paediatric Trauma Essay Example | Topics and Well Written Essays - 2000 words

Paediatric Trauma - Essay Example Details of the case are presented in this paper as well as the proper medical management of this case base on the paramedical standard of paediatric trauma protocol and other reliable sources, utilising theories and related literatures significant in this study to convey a realistic and valid approach of management. Description of the Scene At 08:49, an accident was reported involving a 9-year-old boy, named Martin, who has been hit by a car travelling approximately 30 kilometres an hour whilst crossing the road on his bicycle. On the scene, the boy is conscious with some respiratory distress 10 minutes after the incident. His left leg appears to be deformed, more likely associated to fractured tibia, fibula, or both. A man holding the boy’s helmeted head, maintaining spinal alignment and who claimed that he is the driver of the car and a trained first aider, said that Martin complained about tightness in his chest and that he has asthma. Paediatric Trauma To start with, it is important to emphasize that the term â€Å"trauma† is being used in this study according to its medical definition as a serious or critical bodily injury, wound or shock (MedicineNet.com, 2011). Trauma is the leading cause of childhood mortality because after the age of 5 years, rapid neuromuscular development, frequent social interactions with other children, wider range of activities, and less parental supervision predisposes them to greater risk. Paediatric trauma is more difficult to handle than in adults as presented in Staheli’s (2003, pp. 105-107) ‘Pediatric Orthopaedic Secrets’ book of the contributions made by Dr. Cummings about child versus adult trauma management: Details about the child’s injury will be difficult to gather if the trauma was not witnessed by an adult; assessment is difficult to establish as children frequently resist examination. Trauma causes more death and injury in children than in adults and unintentional injury caus ing blunt trauma is more common to children. Between 30% to 70% of trauma deaths in children are due to head injury because anatomically, children’s head is larger in proportion to the body than in adults and their neck muscles are weaker, thus the cervical spine is more susceptible to injury; their skulls are thinner, and scalps are more vascular than those of adults. Children’s liver and spleen are larger relative to their size than in adults, making them prone to hemorrhage due to intra-abdominal injury. Children are more prone to hypothermia due to higher body surface-to-mass ratio, their skin is thinner and they have smaller stored fats compared to adults. Children cannot be relied on when it comes to immobilization required in fracture healing during recovery as they tend to resume normal activity (if unattended) once they do not feel pain anymore. Assessment of Paediatric Trauma At the moment the health care provider reaches the scene, it is a standard operating procedure that he/she will introduce himself/herself in the crowd while conducting the primary survey and clearing the site for potential harm. Assessment of paediatric trauma in pre-hospital setting should be done rapidly to make sure that the injured child will be transported right away to the nearest facility where in-depth assessment, management, and treatment shall be given. In the pre-hospital setting, the rapid assessment and preparation includes: (1) minimizing scene time, (2) controlling external bleeding and shock, (3) immobilization, (4) immediate transport to the closest appropriate facility, (5) obtaining information from the handover, (6) assessment for level of consciousness, respiratory rate, systolic blood pressure, revised trauma score, and paediatric trauma score, (7)

What Can We Learn about Managemet Today from Exanining Its Historical Essay

What Can We Learn about Managemet Today from Exanining Its Historical Foundations - Essay Example Scientific management theory or classical management theory has its roots from the beginning and which later give birth to behavioral management theory. In scientific theory the focus is on how to increase productivity and efficiency. In order to find the best way to manage workers or employees managers will apply scientific solutions, such as training employees to a specific task to gain organizational goals. This approach only focuses on achieving results and getting productivity ignoring the motivation and will of the workers (Hartness, 47). The major contribution on this approach was put forward by Frederick Taylor also known as the father of scientific management (Kanigel, 8-9). American engineer named Frederick Winslow Taylor published â€Å"THE PRINCIPLE OF SCIENTIFIC MANAGEMENT â€Å"in 1911 was amongst the first modern management publisher inspired by the rise of industrial age (Beissinger, 19). Taylor believed managing people is a science (Taylor, 12). He tried to make ma nagers think of their employees as specialized replaceable components. He emphasizes on directing employees effectively on work more precisely can give the maximum security and success for employer (Aitken, 32). Behavioral management theory on the other hand focuses on how employees will work and how employers could manage the employees in conducting of work in an organizational setting. Its prime concern is employee behavior and expectations and involves the motivation factor which was ignored by scientific approach. During the development of behavioral management theory, a scientist named Elton Mayo conducted an experiment which proved that increasing human motivation and satisfaction will give good amount of productivity. Elton Mayo’s contributions came as a...Furthermore the essay will elaborate scientific school of thought under the working of Fredrick Winslow Taylor who believes employees are machines. Secondly behavioral school of thought will be discussed as per Elton Mayo’s experiments and findings regarding how productivity can be achieved if workers are inclined towards motivation. The work of Abraham Maslow in hierarchy of needs will be discussed and finally the work done by Sun Tzu in terms of behavioral approach will finally be put into consideration. In section two the essay will highlight the core issue regarding continuities and connections between the different stages of management how they are transformed and modified by different people at different era’s and how they were in practices. Furthermore in section two the essay will discuss a problem that has been dominant throughout and its solution given by Fredrick, Maslow and Mayo. Finally then some suggestion and recommendation will be discussed on how the problem can be solved and rectified and what necessary measures should be taken in order to achieve the desired goals.Productivity has always remained a problem for years and the managers of every era have been focusi ng on different techniques to improve the productivity. Some have worked whereas some have not.

Butterflies, by Ian McEwan

Butterflies, by Ian McEwan In their short stories Butterflies, Lamb to the slaughter, The Whole Towns Sleeping and The Pedestrian, Ian McEwan, Roald Dahl and Ray Bradbury create strong feelings of fear and suspense which ultimately lead to death in each story. Discuss these featured in the short stories we have studied. The story Butterflies by Ian McEwan is about an individual is devoid of all emotion, who is also isolated person (maybe because of his look: his chin and his neck are the same thing and he himself thinks he looks grotesque) he takes a walk. A little girl follows him. It was for him a strange thing because he did not have any friends to talk to. He desperately needed affection and in order to keep her close he buys her a doll and ice cream. He lies her saying that at the canal there are boats and butterflies. So the title butterflies is the lure used to take the little girl at the canal. There he abuses her and then he drowns her. I was particularly attracted to this story because of the unusual yet quite sinister way suspense is built up throughout the text. McEwan achieves this by the use of short descriptive sentences; these slow us down and make us feel as if we are an omnipresent being viewing the story as it unfolds. The action takes place in England on Thursday and Sunday. The next text Lamb To The Slaughter utilises descriptive writing right from the opening to massage our senses to the extent we feel we are woven into the story, our minds eye sees everything that needs to be seen, this helps to clearly portray the obvious use of black humour the image of a merry house wife suddenly smashing a frozen leg of lamb meant for dinner over her husbands head is comical in a sinister way. While in Butterflies the opposite is true. The name of the text, suggests something to do with a lamb being slaughtered with is common practice in farms however the phrase can also mean, unaware of any impending catastrophe which when we read on we find out Mr Maloney wasnt aware of himself being in harms way the same is also true for Miss Maloney wasnt aware she would be committing murder she was said to be in shock, Helped bring her out of shock (141). Roald Dahl almost rewards the reader half way through the story with the meaning of the title which we find out is befitti ng after all. The whole towns sleeping employs the use of subtle irony right from the start the title suggests to us that the town is sleeping which they are, its night time but the lonely one is out and the town is oblivious to this fact, Situational irony is also used the reader would become lead to believe that she might come to harm in the woods and when she gets home she will be safe however this is not the case The final text The Pedestrian, the title isnt very exciting it sounds run of the mill, doing this lures us into a false sense of security we dont expect much however out breath is taken by the beautiful way Ray Bradbury starts the piece and our attention is captured. Like Butterflies the writer wants us to savour each sentence and hang of every word so they slow us down therefore building suspense in The Pedestrian this is done with the use of commas. In all of these stories someone is in danger and in two of the stories someone is killed, the stories have a similar of building tempo and setting the pace, also all stories play to our primal emotions weather its humour in lamb to the slaughter or sadness and confusion in butterflies or fear in the whole towns sleeping. Suspense is something that is built up in all stories, in Lamb to the slaughter suspense is built up in an unusual way first the title invites questions such as, who is being killed and is the story based on a farm doing this feeds out curiosity which in itself starts to build suspense and tension. The text starts off quite unusual fashion for a story of this genre, we would expect a dark, dingy and quite frightening place for a murder to take place in this story quite the opposite is true. The opening paragraph starts of quite poetic, The room was warm and clean, the curtains drawn, the two table lamps alight. Dahl also uses descriptive writing to give us vivid picture of a warm homely setting The room was warm and clean. (137) Everything seems above board until we get to the third sentence, the empty chair (137) the use of the word empty suggests that something is missing and in turn that begs the question why is it empty, The Pedestrian uses this also along with the metaphor empty riverbed, The car moved down the empty riverbed streets and off away, leaving the empty streets with the empty sidewalks, and no sound and no motion all the rest of the chill November night. This feeds out curiosity and adds to the suspense. The metaphor helps put across how bare the riverbed was so we have a clear image in our minds. The unusual thing about Lamb to the Slaughter is that the murder happens at the middle there is no time to wonder how the murder will be committed since we dont suspect anything. The reader is taken by surprise much as Patrick and Mary Maloney. The small table over turning helped bring her out of the shock. (141) Mary Maloney and the reader are in shock until we find out she killed him, she didnt mean to kill him it was done out of rage therefore she says So Ive killed him. (141) After the murder scene the reader follows Mrs Maloney around in her meticulous steps to cover her crime. One can wonder how such a sentimental and docile woman can turn out to be so calculating and cool-headed. When the police arrive, we wait for the moment Mrs Maloney gets caught this time never arrives There is an unfulfilled expectation. Butterflies short, snappy sentences to set the tempo which enables the reader to race ahead so we feel their heart is beating in time with the action, Towards midday I decided on a walk. I stood outside the house, hesitating (61). This is in direct contrast with The Pedestrian which opens with a long sentence which slows things down. To enter out into that silence that was the city at eight oclock of a misty evening in November, to put your feet upon that buckling concrete walk, to step over grassy seams and make your way, hands in pockets, through the silences, that was what Mr Leonard Mead most dearly loved to do. This sets a slow tempo so we can really absorb what we are being told and actually feel we are with Mr Mead. Do The whole towns sleeping utilizes the use of conversation to move the story along as well as short and long sentences to set the tempo and pace, we are drawn to the story because of the simple conversations we can empathize with. The theme of fear is something that is rampant in most of the stories, butterflies starts of quite disturbingly, I saw my first corpse on Thursday. This doesnt seem to frighten or upset the boy; he seems to be detached from his emotions unlike Lavinia Nebbs who is so overwhelmed with fear of the unknown she doesnt have the will power to turn round. Dont turn, dont look if you see him, youll not be able to move! Youll be frightened, youll freeze! Just run, run,run! In Lamb to the slaughter instead of being fearful Mary seems to be relishing the moment, she it literally getting away with murder, Mary Maloney began to giggle. While in The Pedestrian and the whole towns sleeping, everyone is fearful in the towns expect the people in actual danger. The way relationships are portrayed differs from story to story Mary Maloney was in what she thought was a loving relationship she loved her husband dearly, Now and again she would glance up at the clock, but without anxiety, merely to please herself with the thought that each minute gone by made it nearer the time when he would come. (137) Mary and her husband had what seemed to be the perfect relationship everything seemed above board, however when she got the news she lost her rationally this could be due to the fact she was pregnant and hormonal for this was her sixth month with child. (137) the strange thing considering it seemed to be done in rage she didnt show any remorse or sorrow, All right, she told herself. So Ive killed him. (141) that could possibly beg the question did she know beforehand did she contemplate the killing before hand, was she acting, does she have split personalities, we know she is calculated and calm under pressure and good at acting when talking to th e shopkeeper after killing her husband she states Patricks decided hes tired and doesnt want to eat out, you know, and now hes caught me without vegetables in the house. When she already knows hes dead and she is the murderer. However know the main character in Butterflies we know is a recluse, who found his mother grotesque he also believes because his chin and neck are close together people see him as untrustworthy he doesnt seem to trust anyone. My chin and me neck are the same thing and it breeds distrust My mother was like that too, only after I had left home did I find her grotesque. She died last year. His mother was also seen as a recluse, he says, Each year she went to Littlehampton and sat on a deckchair by herself, facing out to sea. We get the feeling he wasnt brought up in a loving household and that he didnt have much of a relationship with his mother, which could have lead him to become unsociable and lonely. Although we know he yearns to have some companions when walking in the street a football kicked by a group of boys came his way he stopped the ball with his foot and kicked it back, such a simple gesture caused him to because overwhelmed and giddy I could feel the thumping of my pulse in my temples. Such opportunities are rare for me. I do not meet many people.Also at the start of the text when a simple conversation arose with him and Charlie he fumbled this furthermore stresses the point he was not used to talking to people. When he and Jane converse he immediately realizes he wants her as a friend as she shows genuine interest and curiosity in him, I felt pleased that she was genuinely curious about me, and I was attracted to her. I wanted her to be my friend. The reader isnt alarmed by this as we think he is being friendly but little do we know the wheels have been set in motion for what happens next. After deceiving the girl under the false pretence of seeing butterflies, the disturbed individual wets his finger and wipes the ice cream off from around her mouth and puts the finger back into his mouth and tastes it. While doing this we find out, he has never touched another persons lips and he is excited and seems to be in orgasmic state, I had never touched another persons lips before nor had I experienced this kind of pleasure. It rose painfully from my groin to my chest.'(70) While doing this I believe he was engulfed with pleasure and didnt realize he was pressing so hard on Janes face she tells hi m, You pressed to hard. (70). All throughout the text I is used allot this suggests loneliness. Lavina Nebbs is seen as a father figure she is confident on the surface when around friends they look up to her she doesnt want to show weakness she has a strong relationship with her friends however when alone and going through the ravine she turns into a nervous shell of her former self, Oh god! God, please, please let me get up the hill! (55) Where as Mr Mead from The Pedestrian is calm when he is confronted and even protests his arrest. The whole towns sleeping starts off with the use of descriptive writing to set the scene, It was a warm summers night in the middle of Illinois (37) this gives us an image of brightness cosiness even. Which is the same way Roald Dahl starts Lamb to the slaughter, The room was warm and clean. (137) both authors use this to good effect so the reader is the ability to more clearly visualize what is being described. This subtle use of this type of writing shows itself again throughout The whole towns sleeping in the form of metaphors, Lavina felt the warm breath of the summer night shimmering off the oven-baked sidewalk. We really get a sense that it was a hot day this plays with our senses in the same way in The Pedestrian plays with our sense of smell It smelled of riveted steel (2) and in butterflies a simile is used to the same effect. Like a whippet. (63) The Whole Towns Sleeping. Ray Bradbury uses recreation to create interest to the story In the downtown drugstore, fans whispered in the high ceiling air He also uses metaphors and similes such as There were two moons: a clock moon with four faces The main character in Butterflies isnt really creative we only get one reference to some sort of inner life. Where he makes reference to an artist prominent in the 1920s, she was beautiful in a strange almost sinister way, like a girl in a Modigliani painting. (65) His flat narrative description exactly matches the flatness of his personality, I crossed over and looked at the cars engine, although it meant nothing to me. Another interesting thing about this character is that he never dwells on the fact of death he bounces between subjects. I believe his life experience had an impact on the development of his personality. Like Me Mead in The Pedestrian he is a loner and they both go on walks also the main character in Butterflies has a skewed view on the world whereas in the pedestrian the world is distorted it isnt just in Mr Meads imagination. In all stories there is only really one main character. Mary Maloney is a happy, contented woman who feels secure confident and fulfilled with her life. Roald Dahl describes, There was a slow smiling air about her, and about everything she did. (137) she is loving and the perfect house something that was normal in the time short story was written in 1953. She is a very organised individual she has a routine she does daily, she takes solace in it she believes that if that routine exists, all must be well. She also is calm, collected and rational she methodically gets rid of the evidence, Mary ultimately uses the same means of control over the investigating officers that she had used with Patrick: food, drink, and the illusion of uncomprehending innocence. That being said after all she didnt mean to kill her husband when she realizes what she has done, Mary is willing to accept the consequences, but, she worries about the child: As the wife of a detective, she knew quite well what the penalty would be. That was fine. It made no difference to her. In fact, it would be a relief. On the other hand, what about the child? At the start of the whole towns sleeping Lavina Nebbs is much like Mary Maloney secure and confident she is adamant that fear will not dictate her life, she starts the evening rational and fearless but is reduced to panic on the walk home when she admits her fear. Much like Jane from butterflies she is fearless and outgoing until confronted with fear. Jane who is the girl that was killed in lamb to the slaughter, is a friendly trusting child her parents might have been carefree and unaware that such crimes get committed, most children of that era were told dont talk to strangers or dont walk with strangers however Jane is fearless she is innocent. Jane could be seen as the butterfly a small delicate creature that is gently pushed down the river in the same way a butterfly would fly off into the distance, the word butterflies for a title is very befitting. Lamb to the slaughter is written in the third person in the same way as the whole towns sleeping, the tone of Lamb to the slaughter seems pretty calm it starts of quote poetic each line leading on seamlessly to the next then the tone changes the reader is sped up as Mary Malone executes her plan to get rid of the evidence. The text butterflies is written in the first person the main character is the narrator throughout the story there is a dense and atmospheric tone. Its often left to the reader to give moral significance to events that occur what I found interesting that the story was organized through flashback and the story isnt in chorological order, the story starts of, I saw my first corpse on Thursday (1) when that was the last thing that happened yet mentioned at the start. The narrator also switches from present to past tense. The strength in Butterflies is the writers versatility to be able to move through past and presence tense and to be able to move through different times during the day and to move from subject to subject. However by the end I get a sinking feeling in the pit of my stomach a horrendous crime has been committed and it seems as if the killer will get away, this leaves us wanting and yearning for justice and because unlike Lamb to the slaughter it was a little girl that was abused and killed we feel more hate towards the young man. Lamb to the slaughter the readers get a tale so grotesque, so darkly comic, and so hilarious in some of its incidental details that seem to be something out of a Benny hill show. We feel sympathy towards Mary because her husband broke their wedding vows her act of murder was callus yet unintentional, so we dont yearn for justice although I know a crime has been committed I feel happy to sit back and watch Mary hatch this farfetched comical plan. There could ha ve been emphasis on causing more suspense the detectives could have been more suspicious, however it is a well laid out and quite humorous story and begs the question did the detectives suspect anything after the incident it leaves us with unanswered questions. Roald is also effective at using dialogue to move the story along. Lamb to the slaughter is good at building the tension throughout the story then while Nebbs is in the ravine the excitement builds, when she reaches her house the tempo slows and almost grinds to a halt until she finds out someone is in her house we are left in awe. Ray Bradbury is good at controlling the tempo however at the end of the story I am left feeling unfulfilled I want to know the ending however this may just bring excitement for the sequel and play right into Bradburys hands. In this essay I have discussed the theme of fear and suspense and now different writers portray this and stylistic features used I found the similarity between stories interesting considering there from different authors my favorite story is butterflies I love the simple complexity of it the way the narrator moves from time and place seamlessly. And how all of the stories title have significance to the story and almost give us hints to the plot.