BCH Lab 4 - p 456 2 Unknown 7 11 23pK32 5 UnknownUnknown...

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pK 2 pK 1 Unknown 5 3 2 Unknown 3 1 Unknown 4 7 6 5 4 Unknown 1 Unknown 2 Lab BCH2333 Section: TuA_b (Tuesday Afternoon) LABORATORY 4 Diana Phung Tuesday, February 24 th , 2009 Shaheen Uppal 4918165 Priyen Sondagar 4868085 Purpose:
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Because of their specificity in their functions and reactions with other molecules, proteins are the most important molecules in biochemistry. Proteins are repeating polymers of amino acids, with and N-C-C backbone. There are two specific side groups that are on each amino acid, which are the carboxyl group and the amino group. These groups act as both acids and bases (ampholytes). Because of the differences in composition in the different amino acids, the titration curves and the isoelectric points will also have its own distinct values. In this experiment we are going to observe the differences and study how the amino acids react with different molecules and study how to isolate amino acids, based on their composition. The first experiment is to observe a reaction of glycine, an amino acid. An acid or base titration will alter the pH of an amino acid. In this case, glycine’s pH will be dissolved in a base and titrated with an acid. There will be a loss or gain of electron/protons due to the effects of pH on the ionizable groups on the amino acid. In the second experiment, a mixture of amino acids is going to be separated using ion-exchange chromatography. The mixture contains aspartic acid, glycine and lysine. The ion-exchanger is a cation exchanger, which means that the cations persent in the amino acids will exchange another cation that is bound to the negatively charged immobile phase. In this experiment, the gradient of the increasing buffer pH separates the amino acids from one another. Order of elution is based on separate acid and base characteristics of the amino acids and their isoelectric points. In the third experiment, an SDS-PAGE get electrophoresis is used to separate a mixture of proteins based on the molar weight. The get is composed of b- mercaptoethanol buffer that’s used to reduce the disulphide bridges of the proteins to create monomers from the polymers. SDS detergent is used to arrange the proteins in a linear shape. It does this by binding to the protein, which causes it to fold. The buffer provides mobolity in the PAGE (polyacrylamide gel electrophoresis) to be dependednt on its molar weight. Results and Discussion:
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R1 Table 1. Shows the blank titration and the titration involving glycine in NaOH (0.3 grams of glycine in 20 mL of 0.2 M NaOH) using 2N H 2 SO 4 . BLANK (NaOH) GLYCINE (in NaOH) pH H 2 SO 4 (mL) a Acid Equivalents (L.N)(10 -3 ) b pH Dial (div.) c H 2 SO 4 (mL) a Acid Equivalent s (L.N)b Acid Equivalents (L.N)(10 -3 ) d 11.4 0 0 11.4 0 0 0 0 11.2 0.018 0.036 11.33 9 0.045 0.09 0.054 11 0.035 0.07 11.05 23 0.115 0.23 0.16 10.8 0.053 0.106 10.71 48.2 0.241 0.482 0.376 10.6 0.058 0.116 10.4 84 0.42 0.84 0.724 10.4 0.063 0.126 10.2 113 0.565 1.13 1.004 10.2 0.068 0.136 10 163 0.815 1.63 1.494 10 0.073
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This note was uploaded on 11/13/2010 for the course CHM CHM2132 taught by Professor Goto during the Fall '07 term at University of Ottawa.

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BCH Lab 4 - p 456 2 Unknown 7 11 23pK32 5 UnknownUnknown...

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