Question1 Lactate dehydrogenase catalyzes the following reaction: > lactate + NA

Question

Question1 Lactate dehydrogenase catalyzes the following reaction: > lactate + NAD pyruvate + NADH We will discuss this enzym reaction that it catalyzes. e in detail on class, but for now all you need to know is the overall The following questions are not directly related to material covered in class. However, solving these problems requires only simple algebra and no specialist knowledge of biochemistry. The assay method described is used very frequently in biochemistry labs In a spectrophotometer, NADH absorbs light at 340 nm, NAD does not. The Beer-Lambert Law relates the absorbance of a solution to its concentration, and states that A = .c.L Where A is absorbance, is the molar extinction coefficient, c is concentration, and L is the path length of the solution in the spectrophotometer. In all the calculations that follow assume L = 1 (ie all measurements were made in a cuvette with a 1 cm path length), so L can effectively be disregarded (although not its unit) The molar extinction coefficient for NADH is 6,300 litres/mol cm [or (mo/litre)1 cm or M cm]. The molar extinction coefficient is the absorbance of a 1 molar solution in a 1 cm path length cuvette. The molecular weight of NADH is 663.4 In a 1 ml cuvette, you mix 0.1 ml of 2.5 mM NADH, 0.1 ml of 0.2 M pyruvate, 0.7 ml of a suitable buffer, and 0.1 ml of a solution of purified lactate dehydrogenase. You mix, place the cuvette in a spectrophotometer (with a chart recorder attached) and follow the absorbance at 340 nm. You measure a rate of decrease of absorbance (AA) of 0.51 min1. You are told that the lactate dehydrogenase solution you used in the assay contained 8.17 mg of protein per ml.

Explanation / Answer

Specific activity lactate dehydrogenase is the amount of enzyme needed to form 1umoleof NADH/min.

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