9. Suppose you have just purified an enzyme (lactase dehydrogenase - LDH) and wa
ID: 37746 • Letter: 9
Question
9. Suppose you have just purified an enzyme (lactase dehydrogenase - LDH) and want to check its activity. You know that in a pH 9 Tris buffer that LDH catalyzes the following reaction: lactate + NAD LDH pyruvate + NADH You are not aware of a simple colorimetric assay to monitor this reaction. You performed two wavelength scans, one on purified NAD and one on purified NADH: You decide to use the spectrophotometer to monitor the LDH activity. (3pts) List all the components that should be in the ''blank'' cuvette for this assay. (1 p1) Prior to adding LDH how much pyruvate should be present in the assay tube. (2pts) What is the best wavelength used to measure the LDH activity? (4pts) After adding LDH to the assay tube, you begin monitoring the reaction. What would you expect to happen to the absorbance while measuring at the ''best'' wavelength? (2pts) What is the second best wavelength that could be used to measure the LDH activity? (3pts) Explain why this wavelength should not be the first choice used?Explanation / Answer
blank- LDH in tris buffer pH=9
450nm
2nd best 492 nm
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