2. You are studying a dehydrogenase enzyme that catalyzes the formation of NADH

Question

2. You are studying a dehydrogenase enzyme that catalyzes the formation of NADH from NAD Subsequent studies determined that NADH will absorb at 340 nm while NAD will not. The extinction coefficient for NADH is 6220 MT cm The reaction volume is 750 L total and the molecular weight of the substrate is 223 g/mol. You use 20 uL of enzyme from an initial concentration of 20 mg/mL for everyassay. The molecular weight of the enzyme was found to be 26 kDa a. What volume of a 3.4 mg/mL initial stock solution for the substrate would be needed to measure a final concentration of 200 M for one reaction? (5 points)

Explanation / Answer

The concentration required = 200X 10^-6 Molar

We have concentration = 3.4 mg /mL = 3.4 g/L

Molecular weight of substrate = 223g/mole

so the inital molar concentration = 3.4 /223 = 0.015 molar

We need a volume of 750 micro liter of 200 X10^-6 molar solution

so the volume of stock required can be calcualted as

M1V1 = M2V2

0.015 X V1 = 200 X10^-6 X 750 microliter

V1 = 100 microliter

So we need 100 microliter of stock solution to preapre the desired solution for 750 microliter of volume

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