Microsoft word - enzyme engineering & technology

IB 243 – ENZYME ENGINEERING & TECHNOLOGY a. Enzymes used for the manufacture of fructose syrup from b. A recombinant enzyme approved for use in the dairy products. 2. Explain with an example indirect assay of an enzymatic reaction. 3. Write down 2 methods of separation of proteins from low molecular 4. Sucrose + H2O ---> glucose + fructose The equilibrium constant for the conversion of the disaccharide sucrose to the simple sugars glucose and fructose is 140,000. What can you conclude from the above? (a) Its a closed system (b) It never reaches equilibrium (c) It is spontaneous, starting with sucrose (d) At equilibrium, the concentration of sucrose is much higher than the concentration of glucose and fructose. 5. What are Biosensors? Give an example 6. Explain Thiele Modulus. Write down the expression for Thiele modulus for a first order reaction, spherical geometry. 7. List any 2 downstream processing steps involved in the extraction of 8. When alcohol dehydrogenase was immobilized on polyacrylamide, Km for Butanol was 0.1 mM. When the enzyme was immobilized on a polymer of methacrylate and acrylamide, Km for Butanol was found to be 0.025 mM. Explain the above (Assume all other conditions to be identical for both the cases) and explain how one will evaluate the true kinetic parameters. 10. Methanol is a poison not because of what it does to the body itself but because the enzyme alcohol dehydrogenase oxidizes it to formaldehyde, which is a potent poison. A treatment of methanol poisoning is to give the patient ethanol (CH3CH2OH). Why is this effective? (Question No. 11 is Compulsory) (5 x 16 = 80 Marks) 11. The kinetics of an enzyme were analyzed in the absence and presence a) What type of inhibitors are A and B: Competitive or NonCompetitive? (Use your graphs to answer this question). b) In addition to calculating Km and V (mazimum velocity) in the absence of inhibitors, calculate the Ki constants for the inhibitors A and B. 12. Discuss the application of enzymes in the food and detergent industry Outline the method of manufacture of High Fructose corn syrup and explain the steps involved. 13. Derive an expression for the concentration profile of a substrate following first order kinetics and spherical geometry considering internal mass transfer. Explain the different methods of immobilization of enzymes, discussing the advantages and disadvantages. 14. Penicillin is hydrolyzed and thereby rendered inactive by penillicinase (also known as β-lactamase), an enzyme present in some resistant strains of bacteria. The molar mass of this enzyme is 29,000 g/mole. The amount of penicillin hydrolyzed in 1 minute in a 10 mL solution containing 10-9 g of purified enzyme was measured as function of the concentration of penicillin (in µmol/L). Assume the concentration of penicillin does not change appreciably during the assay. a.) Plot 1/rate vs. 1/[S] for these data. b.) What is KM? c.) What is Vmax? d.) What is the turnover number? a) Rearrange the Michaelis – Menton equation to give Eadie-Hofstee b) What is the significance of the i) slope ii) Vertical intercept iii) c) The reciprocal plot of a Bi-Bi reaction with ping pong mechanism for varying fixed substrate concentration generates parallel lines. Explain the above with the equation. Also provide a theoretical explanation. 15. a) Define Damkholer Number, External effectiveness factor. b) Derive the relationship between External effectives factor ηe and Damkholer number An enzyme is used to produce a compound used in manufacture of sunscreen lotion. Vmax for the enzyme is 2.5 mmol m-3 s-1; Km = 8.9 mM. The initial concentration of substrate is 12 mM. (a) Find the batch time required to achieve 90% substrate (b) Assume the enzyme deactivates with half-life of 4.4 hours. Calculate the batch time for 90% substrate conversion and compare it with the time required without deactivation.

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