Key HW 3 Fall, 2009

Key HW 3 Fall, 2009 - 1 Biological Sciences 102 Fall 2009 K...

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1 Biological Sciences 102 Answer Key Fall, 2009 K. Hilt Homework #3 Assigned text problems: Biochemical Calculations (Segel) 4-4 on pp. 223-224 and 4-9 on pp. 237-238 pp. 319 #2, 3, 6, 10, and 12. You need to be able to plot and interpret Lineweaver-Burk graphs. 1. Enzyme Z catalyzes the reaction: A + NAD + B + NADH + H + Besides following the [H + ] using a pH meter, the reaction can be monitored spectroscopically at 340 nm. This is the wavelength at which NADH absorbs light. Some of the first enzymes that were worked on in biochemistry were dehydrogenases, like our enzyme Z, because of this advantage. Radioisotopes were not yet available. a) Diagram what the kinetic assay results would be if the reaction was measured from left to right. What would it be if the assay was measured from right to left? [NADH] left to right (red line) r i g h t t o l e f t (green line) Time b) Describe how you would accurately determine enzyme Z's K m for NAD + . Describe the experimental set-up and the analysis of data. Assay a series of tubes where the concentrations of buffer, compound A, and enzyme Z were constant. Vary the concentration of NAD + . Make a Lineweaver-Burk plot and determine the K m from the x-intercept: 1 / v o 1/[S] c) Imagine that twenty I.U.’s of enzyme Z were catalyzing the above reaction for one minute, under V max conditions, in a 3.00 ml assay volume. The assay is buffered with 20 mM phosphate buffer, pH 7.60. What will the pH be at the end of that one minute? pH 7.0 Use two Henderson-Hasselbalch equations. This question is very similar to questions #40 and #41 in Biochemical Calculations (Segel), on page 93, which were assigned in homework #1.
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This note was uploaded on 12/03/2009 for the course ENT 100 taught by Professor Kimsey,r during the Fall '08 term at UC Davis.

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Key HW 3 Fall, 2009 - 1 Biological Sciences 102 Fall 2009 K...

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