6. You are given a mixture of proteins that you analyze by standard 2-D electrop
ID: 187065 • Letter: 6
Question
6. You are given a mixture of proteins that you analyze by standard 2-D electrophoresis, with the following results for isoelectric points and apparent molecular weights: protein A (Mr 110,400; pl 4.60), protein B (Mr 10,100; pl 6.93), protein C (Mr 65,200; pI 7.84), and protein D (M 25,000; pI 8.15) a. After the 2-D separation, which protein will be in the upper left quadrant of the gel. given that e cathode-proximal end of the isoelectric focusing gel was on the right side of the 2-D gel? plain why, citing the physical basis of each separation step as part of your answer drawa me of the 2D-gel indicating where the electrodes positive and negative- are located in each step of the 2D gel electrophoresis) b. Which of the four proteins is the most acidie? Explain why. c. Upon gel filtration of the mixture, only 3 peaks are seen. Their molecular masses, estimated using a calibration curve with globular proteins of know Mr, correspond to 160 kDa, 65 kDa, and 10 kDa. What can you conclude if you assume that they all behave as globular proteins? Explain why d. Protein C is phosphorylated at two Tyrosine residues. After treatment of the proteins with a phosphatase, would you expect that its pls would be shifted to higher or lower values? Explain why If a mixture of B and C is subjected to carboxymethyl (CM, pKa 4.76) matrix chromatography. what optimal pH range should you use for the buffer and in what order will the proteins emerge from the column? Explain why e. If the mixture of B and C is subjected to salting in with KCI at pH 6.8, in what order would you expect the proteins to be solubilized? Explain why. f.Explanation / Answer
Answer b:- The most acidic proteins are as aspartic acid, glutamic acid, asparagine and glutamine. These are acidic in nature. These are acidic in nature because of extra carboxylic group. And are known to be hydrophilic in nature and are polar. They are found to be on a protein molecule when they come in contact with water.
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