2024 Engineering Engineering / Homework 2 Due: April 18, 2024

Solve the following problems. Be sure to draw the graph using a computer and write down the units of your answers.

1. The following data were recorded for the enzyme-catalyzed reaction S→P.

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S (M)                     V (nM/min)

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6.25 x 10-6                    15

7.50 x 10-5                    56.25

1.00 x 10-4                    60

1.00 x 10-3                    74.9

1.00 x 10-2                    75

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1) Estimate Km and Vmax.

2) What would V be at S=2.5 x 10-5 ?

2. The following data were obtained for an enzyme that catalyzes the reaction

S→P. Estimate Km and Vmax.

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S (M)                     V (nM/min)

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8.33 x 10-6                    13.8

1.25 x 10-5                    19

2.00 x 10-5                    26.7

3.33 x 10-5                    36.3

5.00 x 10-5                    44.4

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3. Determine the Michaelis-Menten equation for the following reaction:

urea + urease = [urea․urease] → 2 NH3 + CO2 + urease

The rate of equation is given as a function of urea concentration in the         following table.

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Curea (kmol/m3)              V (kmol/m3s)

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0.2                        1.08

0.02                       0.55

0.01                       0.38

0.005                      0.2

0.002                      0.09

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4. The following data are oxygen uptake rate of bakers' yeast for enzymatic reaction in a particular medium at 25℃ and various oxygen partial pressure were obtained. Determine kinetic parameters.

PO2 (mmHg)	QO2 (M/h)
0	0
0.5	23.5
1.0	33.0
1.5	37.5
2.5	42.0
3.5	43.0
5.0	43.0

5. We can directly determine kinetic parameters (Vmax and Km) in terms of reaction time (t) versus substrate concentration (S0 and S). Calculate the Vmax and Km using the following data. You may use a semi-log plots for solving this problem.

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Time (min)           S (mol/liter)

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0                     0.1

10                     0.089

20                     0.079

50                     0.053

100                     0.025

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6. Malate dehydrogenase catalyses the reaction:

(S)-malate + NAD+ = oxaloacetate + NADH+H+

The rate of the forward reaction was investigated in the presence of saturating concentrations of malate and a fixed concentration of enzyme. The following results were obtained. Calculate the kinetic parameters (Km and Vmax) in this reaction.

NAD+ concentration (mmol/l)	Absorbance at 300 nm (A300) at time t (min) x 102
	t=0.5	t=1	t=1.5	t=2	t=2.5	t=3
1.5	3.3	5.6	7.9	10.2	12.2	13.8
2	3.6	6.3	8.9	11.6	13.8	15.4
2.5	4.0	6.9	9.9	12.8	15.0	16.8
3.33	4.3	7.5	10.8	14.0	16.3	17.5
5	4.7	8.4	12.1	15.8	17.7	18.4
10	5.2	9.5	13.7	18.0	19.2	20.0

[Hints] You must first determine the reaction velocity for each cosubstrate concentration (NAD+).

(Example) For 1.5 nmol/l NAD+

A300

Slope=reaction                   velocity

time (min)