Typical Examination Questions

You will be required to answer any three questions in two hours

1. How does the effect of substrate concentration on the rate of an enzyme catalyzed reversible reaction differ from that on non-reversible reactions. (25%)

The kinetic constants and starting conditions for the reversible glucose isomerase reaction

Glucose ® Fructose

were determined to be:

K_m (glucose) = 100 mM = K_m^S

K_m (fructose) = 120 mM = K_m^P

V_max (glucose ® fructose) = 10 mmol min^-1 = V^f

Equilibrium constant = 1.1 = K_eq

Glucose concentration = 1 M

Fructose concentration = 0 [S_ is the substrate concentration at equilibrium

Given that the reaction velocity v = V[S#]/(

where V = ((Keq + 1)/Keq) (KmP

and K + (KmS KmP + (KmP +

Calculate the rate of reaction when the fructose concentration is 0.5 M. (75%)

2. Write an illustrated account of the use of enzymes in the laundry and clothing industries.

3. Employed by a small biotechnology company, you have been given the task of developing a biosensor for lactic acid in blood for use in assessing athletic performance. Describe how you would set about your task. Include within your answer (i) the type or types of biosensor that you might investigate, (ii) the mechanism for the biosensor response, and (iii) the likely advantages and disadvantages of analyses using your biosensor(s) relative to analyses by other methods.

(Do not answer this question with a general essay on biosensors)

4. The relationship giving the time taken (t) for an enzyme-catalyzed reaction to achieve a particular fractional conversion is provided below:

V_max.t = [S]₀ X - K_mLn(1 - X)

Explain this relationship paying particular attention to the meaning of the terms and the reason why there are two parts to the right hand side of the equation. (50%)

A batch reactor containing the enzyme lactase is used to hydrolyse a 5 % w/v solution of lactose (molecular weight 342) into a mixture of glucose and galactose. If the K_m and V_max of the enzyme used are 1 mM and 600 mol min^-1 l^-1respectively, calculate the time required to achieve 80% conversion. (50%)

5. Write an essay about the use of carbohydrate hydrolases by industry.

6. Given that the forward rate (v) of a reversible reaction is given by:

v = V[S#]/(

and the kinetic constants and starting conditions for a glucose isomerase reaction were determined to be:

V = 20 mmol min^-1 litre^-1

K = 2 M

K_eq = 1.1

Glucose concentration = 1 M

Fructose concentration = 0 M

Reactor volume = 2000 litres

What is the meaning of [S^#]? (4 marks)

What is the productivity rate when the reaction is 99% complete? (16 marks)

7. Write an illustrated account of EITHER the analytical uses of enzymes and cells OR the industrial uses of immobilised biocatalysts.

8. The relationship between the time of reaction and the fractional conversion (X) of a batch reactor is given by

V_max.t = [S]₀ X - K_mLn(1 - X)

(a) What is the meaning of X. (4 marks)

(b) A 2000 litre batch reactor containing the enzyme eurekase is used to convert a 10% w/v solution of a medium value natural product (molecular weight 355, £1000 Kg^-1) into the high value pharmaceutical curitall (£10 g^-1). If the K_m and V_max of the enzyme used are 10 mM and 1 mmol min^-1 litre^-1 respectively, draw a graph showing how the value of the contents of the reactor varies with the operating time. If the overheads of running the reactor are £1000 day^-1 and the downstream processing costs are insignificant, for how long should the batch reactor operate.

9. Write an essay on the enzymic processing of starch.

10. Write an illustrated account of EITHER the pharmaceutical use of enzymes OR enzyme production.

11. Write an essay about the use of enzymes in the food industry.

12. (a) Briefly describe how inhibitors may affect enzyme catalysed reactions. (5 marks)

(b) The initial velocities of an enzyme-catalysed reaction were measured in the presence and absence of an inhibitor. The data for these measurements are given below.

[S] (mM)	Initial velocity (mol min-1)
- Inhibitor	+ Inhibitor (10 mM)

Graphically determine the type of inhibition (15 marks)

13. Write an illustrated account of the use of enzymes in EITHER the leather and clothing industries OR the dairy industry.

14. Write an essay about the industrial production of enzymes.

15. Describe the different types of kinetic mechanisms that may be found in two substrate two product reactions. (12 marks)

An esterase may also act in transesterification reactions as shown below:

C₂H₅OOCCH₃ + C₂H₅COOH C₂H₅OOCC₂H₅ + CH₃COOH
ethyl acetate + propionate ethyl propionate + acetate

It is found that acetate competitively inhibits the forward reaction with respect to propionate but shows mixed inhibition with respect to ethyl acetate, Also ethyl propionate competitively inhibits the forward reaction with respect to ethyl acetate but shows mixed inhibition with respect to propionate. Explain the likely mechanism for this reaction. (8 marks)

16. Write an illustrated essay on enzyme mechanisms

17. Describe in detail the mechanisms by which biosensors may make use of oxidoreductase enzymes.

18. Describe the use of enzymes in the starch industry

19. The relationship giving the time taken (t) for an enzyme-catalysed reaction to achieve a particular fractional conversion is provided below:

t = (K_m/V_max).{ X- Ln(1 - X)}

(a) Explain this relationship paying particular attention to the meaning of the terms and how the equation would differ if the reaction ran at V_max throughout. (10 marks)

(b) A batch reactor containing the enzyme invertase is used to hydrolyse a 2 % w/v solution of sucrose (molecular weight 342) into a mixture of glucose and fructose. If the V_max and K_m of the enzyme used are 2 mmol min^-1 l^-1 and 5 mM respectively, determine, using graphical means, the % conversion after 40 minutes. (10 marks)

20. (a) Explain how enzymes lower the activation energy for a reaction. (12 marks)

(b) Given the following data on an enzyme catalysed reaction, calculate the activation energy (R = 8.315 J.mol^-1.deg^{- 1}). (8 marks)

Temperature (°C)
Vmax (mol/min)	0.74	1.27	2.11	3.40	1.91

21. Write an illustrated essay on allostery

22. Describe in detail the use of the coenzymes in the mechanism of pyruvate dehydrogenase

23. Describe the use of enzymes in those parts of the non-corn-starch food industry (i.e. write about the food applications of enzymes but do NOT include the production of dextrin or glucose from corn starch).

24. Determine the kinetic parameters and comment on the following data which was obtained for the inhibition of an enzyme..

Substrate concentration (mM)	Uninhibited rate (mol/min)	+ 1 mM Inhibitor (mol/min)
	0.86	0.35
	1.33	0.63
	1.64	0.86
	1.85	1.04
	2.00	1.20

25. Describe, with the aid of the mechanism(s) of an actual enzyme(s), what is meant by the term 'Ping-Pong mechanism'. (10 marks)

How can the use of inhibitors confirm that the mechanism is 'Ping-Pong'? (10 marks)

26. Write an illustrated essay on the mechanisms of reactions involving coenzymes.

27. Describe in detail the mechanisms by which biosensors may make use of light (optical biosensors).

28. Write an essay on the enzymes involved in the production of glucose from starch.

29. Giving examples, describe how enzymes may be used as analytical reagents in solution.

30. Write an illustrated essay on the use of enzymes by the clothing and laundry industries.

31. Describe in detail the mode of action of the coenzymes in the mechanism of pyruvate dehydrogenase.

32. Explain the differences in the following data which was obtained for an enzyme- catalysed reaction, using the same starting material.

Time (hrs)	Total product formed (moles) at 50 °C	Total product formed (moles) at 60 °C
	0.91	1.53
	1.69	2.47
	2.34	3.03
	3.36	3.58
	4.25	3.81
	4.83	3.87

What conclusions can you draw about the economics of the process if you take into account possible running costs?

33. (a) Why is the direct linear plot preferred over the double reciprocal plot in the determination of the K_m and V_max of an enzyme? (8 marks)

(b) Using the direct linear plot, determine the K_m and V_max of an enzyme from the following data. (12 marks)

Substrate concentration (mM)	0.1	0.2	0.4	0.8	2.0	4.0
Initial rate of reaction (mol/min)	0.52	0.88	1.27	1.76	2.45	2.56

34. Give an overview to the analytical applications of enzymes, to include their use in biosensors and as analytical reagents. Give specific examples stating how they work and why they are used in preference to other analytical methods.

Either

Compare the kinetics of reversible and non-reversible enzyme-catalysed reactions. (10 marks)

Show how the kinetic equations for a reversible reaction may be developed, stating the assumptions that need to be made. (A full derivation is NOT required) (10 marks)

Or

Compare the kinetics of inhibited reactions. (10 marks)

Show how the kinetic equations for inhibited reactions may be developed. (10 marks)

36. (a) Explain what is meant by the 'productivity' of an enzyme in a process explaining the factors that might limit it. (8 marks)

(b) Given the expression

(V_max/Vol_S).t = [S]₀ X - K_mLn(1 - X)

determine the percentage hydrolysis of 5 litres of 2 M lactose in 15 hours. (K_m = 2 mM; V_max = 10 mmol/min). (12 marks)

This page was last updated by Martin Chaplin
on 18 January 2002

Proteomics tools for biomedicine (2002): Wasinger, V. C.and Corthals, G. L. (2002). J Chromatography B: Analytical Technologies in the biomedical and life sciences: 771(1-2): 33-48.

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