ExtraCreditHomework2_KEY - Name: KEY EID: Extra Credit...

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EID: Extra Credit Homework #2 Complete questions below and hand in to a TA by the end of class, Wednesday September 7. 1. Protein Purification. Triose phosphate isomerase (or TIM) is a metabolic enzyme that interconverts dihydroxyacetone phosphate and D -glyceraldehyde-3-phosphate, a key reaction in glycolysis. P O O O O OH P O O O O OH O O Dihydroxy acetone phosphate D-glyceraldehyde-3-phosphate a) Saccharomyces cerevisiae (yeast) TIM is a 248 amino acid protein. Estimate its molecular weight. The average molecular weight of an amino acid is 110 Da, so a good approximation would be ~27 kDa. If you calculated it by amino acid sequence, it turns out to be 26.7 kDa (a good applet for calculating molecular weight, isoelectric point, and other useful information for primary sequence http://www.scripps.edu/~cdputnam/protcalc.html if you’re curious). b) If S. cerevisiae TIM with a genetically-engineered, appended hexahistidine affinity tag were over-expressed in Escherichia coli and purified through a nickel affinity column, what would you expect the indicated samples to look like on a SDS-PAGE gel? Sketch the expected protein bands. ~250
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ExtraCreditHomework2_KEY - Name: KEY EID: Extra Credit...

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