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King Saud University College of Science Department of Biochemistry BCH 221 – Enzymology Final Examination 2nd Semester 1419-1420 Name:_____________________________________ No.__________________ Time: 3 hours. Final Marks Section 1 ………… /10 Section 2 ………… /40 Section 3 ………… /10 Section 4 ………… /5 Section 5 ………… /25 Section 6 ………… /10 Total: …. /100 Total: …… /60 Final Marks CAT: _________ /40 Final: _________ /60 Total: _________ /100 Grade: _______ BCH 221 Final Section 1 Answer the following questions as True [T] or False [F]: 1. An enzyme can have different Km for the same substrate. [ ] 2. Eadee-Hofstee plot is obtained when [S]/v is plotted against [S] [ ] 3. Any substance that binds the active site of an enzyme is known as a substrate. [ ] 4. All enzymes have a quarternary structure. [ ] 5. Haloenzymes are conjugated proteins. [ ] 6. Coenzymes are derived from fat soluble vitamins. [ ] 7. Absence or deficiency of melanin due to deficiency of enzyme tyrosinase produces albinism. [ ] 8. pH changes have no effect on the charge on the enzyme. [ ] 9. Enzymes have an ability to take-in or give-out H+ (protons). [ ] 10. The maximum absorption at 280 nm given by enzymes is due to the presence of aromatic amino acids. [ ] 11. At the isoelectric pH the enzyme has no negative or positive charge. [ ] 12. Standard free energy (G°) is the difference in the energy level of transition state and reactants. [ ] According to the international classification of enzymes, enzymes are placed in 6 classes depending on the substrate on which they act. [ ] The following reaction will be catalysed by an enzyme belonging to class 6 of enzyme classification: A - B + C A-C + B [ ] 15. Glucokinase is an enzyme with absolute specificity. [ ] 16. Velocity (v) of the following reaction S P is equal to –d[p] dt t½ is the time in which the concentration of substrate is reduced to half its original concentration. [ ] [ ] 13.. 14. 17. Page 2 of 11 18. BCH 221 Final In the Lineweaver-Burk plot the intercept on the axis on which 1/[S] is plotted is equal to 1/Km. [ ] 19. When v is plotted against v/[S] the slope of the line gives-Km. [ ] 20. If a non-competitive inhibitor is added to reaction E + S P + E, the concentration of free enzyme [EF] is equal to: [ET]-ES]-[EI) [ ] Section 2 In this Section each statement is followed by FOUR related statements which may be all true, all false or a mixture of true and false. Put [T] in front of the true statement and [F] in front of the false one: 1. Different coenzymes are involved in different reactions: (a) (b) (c) (d) 2. Coenzyme A transports acyl groups. FAD transports CO2. All carboxylases used biotin. NAD+ is involved in H+ transport. [ [ [ [ ] ] ] ] [ [ [ [ ] ] ] ] In the following curve: (a) (b) (c) (d) [S] represents molar concentration of substrate. Point (b) is called the initial velocity. Point (c) is due to mixed order reaction. Point (a) occurs as the rate is independent of substrate concentration. Page 3 of 11 BCH 221 Final 3. Cofactors: (a) (b) (c) (d) 4. ] ] ] [ ] Tyrosine. Histidine. Phenylalanine. Threonine. [ [ [ [ ] ] ] ] [ [ [ [ ] ] ] ] [ [ [ [ ] ] ] ] [ [ ] ] [ ] [ ] The following enzymes belong to class I of enzyme classification: (a) (b) (c) (d) 6. [ [ [ Enzymes give a maximum absorption at 280nm. This is due to the presence of: (a) (b) (c) (d) 5. are always organic molecules. combine with haloenzyme to form apoenzyme. are generally stable to heat. are essential for the activity of the enzyme which requires the specific cofactor. Oxygenases. Aldolases. Carboxylases. Reductases. The following reaction: Glucose + ATP Glucose-6-phosphate + ADP is brought about by enzymes (a) (b) (c) (d) 7 belonging to class 2 of enzyme classification. generally known as kinases. lyases. synthetases. The specific activity of an enzyme was 240 and you had 2 mg of this enzyme: (a) (b) (c) (d) Each mg has 240 units of enzyme activity. Each mg has 120 units of enzyme activity. You purified this enzyme more and the specific activity become 360. This means that the amount of protein decreased. During heating the specific activity decreased as protein concentration increased. Page 4 of 11 BCH 221 Final 8. Spectrophotometer can be used to study the following enzymic reactions: (a) (b) (c) (d) 9. [ [ [ [ ] ] ] ] is a constant. Shows the affinity between enzyme and substrate. is decreased by presence of competitive inhibitors. = k-1+k+2. k+1 [ [ [ [ ] ] ] ] [ [ [ [ ] ] ] ] Km: (a) (b) (c) (d) 10. Fumerate malate. G-6-P+NADP 6-phospho-gluconolactone + NADPH Phosphoenol pyruvate + ADP Pyruvate + ATP. + + Dehydrogenases using NAD or FAD as H -ion acceptor. If we plot 1/v vs [I] at two different substrates concentration, we obtained the following graph: (a) (b) (c) (d) This indicates competitive inhibition. This is the Lineweaver and Burk plot. Point (c) on the graph gives -Ki. Point (b) gives - 1/km. Page 5 of 11 BCH 221 Final Section 3 In this Section each statement is followed by four related statements, only ONE of which is correct, put [T] in front of the correct answer.: 1. Enzymes: (a) (b) (c) (d) 2. are only coenzymes. are strongly linked to the enzyme protein. may be removed from an enzyme molecule without loss of activity . are only metal ions. [ [ [ [ ] ] ] ] can bind a competitive inhibitor. can be altered without loss of enzyme activity. binds noncompetitive inhibitors. is always hydrophobic. [ [ [ [ ] ] ] ] [ [ [ [ ] ] ] ] When pH of an enzyme catalysed reaction is changed from pH 8.6 to 12.5: (a) (b) (c) (d) 5. ] ] ] ] Active site of an enzyme: (a) (b) (c) (d) 4. [ [ [ [ Prosthetic groups: (a) (b) (c) (d) 3. increase rate of reaction by decreasing Gibbs Free Energy G°. are changed during catalytic reactions. are all simple proteins. do not change K equilibrium of a reaction. the enzyme becomes positively charged. the enzyme precipitates. the rate of reaction increases. the enzyme becomes negatively charged. The following substrate: Page 6 of 11 BCH 221 Final R H2N – C – COOH H is optically active as: (a) (b) (c) (d) 6. it has one –NH2 group. it has one COOH group. it has one R group. it has assymetric C atom. (a) (b) (c) (b) is endothermic. must get energy from another source to proceed. has negative (-) value for G°. has positive (+) value for G°. ] ] ] ] a ligase. a lyase. an isomerase. a transferase. [ [ [ [ ] ] ] ] [ [ [ [ ] ] ] ] [ [ [ [ ] ] ] ] The following coenzymes are not involved in H+ transfer: (a) (b) (c) (d) 9. [ [ [ [ P + 5 Kcal (energy) The number of an enzyme is 5.3.4.7: It is: (a) (b) (c) (d) 8. ] ] ] ] A reaction: A 7 [ [ [ [ FAD+. NADP+ FMN. Coenzyme A. The enzyme protease: (a) (b) (c) (d) has absolute specificity. is a hydrolase. links amino acids to form proteins. belongs to class 2 of the enzyme classification. Page 7 of 11 BCH 221 Final Which of the following methods is most appropriate to study the following reaction: 10. Pyruvate + CO2 Oxaloacetate. Pyruvate carboxylase (a) (b) (c) (d) Electrode method. Fluorescence method. Spectrophotometric method.. Manometric method. [ [ [ [ ] ] ] ] Section 4 Complete the following Table for an enzyme extracted from potatoes: Step Total volume Total units protein/m l Specific activity purification Yield (%) Extract 4520 20450 30 ……… 1 100 (NH4)SO4 Precipitation (4045%) 590 18290 …… 4.2 ……… …….. 370 ……. 1.4 48.0 …….. …….. 100 15400 0.21 ………. ………. ……… DEAE Cellulose (eluate) Affinity chromato-graphy (eluate) Page 8 of 11 BCH 221 Final Section 5 Write short notes on the following: 1. Significance of enzymes in diagnosis. [5 Marks} 2. Enzyme specificity. [5 Marks] Page 9 of 11 BCH 221 Final [5 Marks] 3. What are inhibitors and what are their uses. 4. Propeties of enzymes. [5 Marks] 5. Methods for assaying enzymes. [5 Marks] Section 6 Page 10 of 11 BCH 221 Final Section 6 In an experiment carried out in your BCH 221 laboratory the following initial velocity was obtained when different amounts of substrate were added to a fixed amount of enzyme. The experiment was repeated again in presence of 0.00022M inhibitor. v (µmol/min) S (mmol/l) without I with I 1.0 x 10-1 28 17 1.5 x 10-1 36 23 2.0 x 10-1 43 29 5.0 x 10-1 65 50 7.5 x 10-1 74 61 Using Lineweaver-Burk plot determine Km, Ki and Vmax in presence and absence of inhibitor. What is the type of this inhibitor. Page 11 of 11