Student Number: THE UNIVERSITY OF MANITOBA April 10, 2006, 1:30 AM - 4:30 PM Page 1 (of 4) Biochemistry II Laboratory Section Final Examination

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1 Name: Student Number: April 10, 2006, 1:30 AM - 4:30 PM Page 1 (of 4) Biochemistry II Laboratory Section Final Examination Examiner: Dr. A. Scoot 1. Answer ALL questions in the space provided. 2. The back side of each page may be used for your answer or for preliminary work. 3. Questions to invigilators about the exam will not be answered State which chemicals or products were used in the lab this term for each of the following purposes. N.B. Full names and correct spelling required. A) To initially precipitate the RNA, DNA and protein from liver homogenate. Ans B) To yield a purple complex upon reaction with peptide bonds in the assay to measure protein concentration. Ans C) To form a coloured complex in the assay used to measure RNA concentration. Ans D) To form a coloured complex in the assay used to measure DNA concentration. Ans 4 2. Use a figure to illustrate the staining sequence used in the lab this term for lactate dehydrogenase.

2 April 10, 2006, 1:30 AM - 4:30 PM Page 2 (of 4) Biochemistry II Laboratory Section Final Examination Examiner: Dr. A. Scoot 7 3. Two methods of column chromatography were used to isolate glutamate dehydrogenase from a partially purified liver homogenate. Enzyme activity in 0.1 ml of a 1 in 50 dilution of the homogenate was 4.0 mmoles -ketoglutarate ( -KG)/min. Protein content of 0.5 ml of a 1 in 50 dilution of the homogenate was found to be 4.0 mg. A 5 ml portion was applied to Column A and another 5 ml applied to Column B. Several 10 ml fractions were collected from each column. For Column A the enzyme activity was found in fraction 2 where 0.1 ml of a 1 in 10 dilution produced 5.0 mmoles -KG/min and the protein content of 1.0 ml of a 1 in 10 dilution was found to be 2.5 mg. For Column B the enzyme activity was in fraction 6 where 0.1 ml of the fraction produced 20 mmoles -KG/min and the protein content of a 1.0 ml aliquot was found to be 5.0 mg. (a) Which of the two columns would you chose if you were interested in the best percentage yield of enzyme? Show all calculations and cite values to support your answer (b) Which of the two columns gave the best purification of the enzyme? Show all calculations and cite values to support your answer. (c) Based upon your experience in the biochemistry lab this year briefly describe an assay you could use to detect the presence of this enzyme.

3 April 10, 2006, 1:30 AM - 4:30 PM Page 3 (of 4) Biochemistry II Laboratory Section Final Examination Examiner: Dr. A. Scoot 2 4. In the lab this term barium hydroxide and zinc sulphate were used in the same experiment. Which experiment was this and what was their purpose? 2 5. Potassium hydroxide played an important role in an experiment this term. Which experiment was this and what was the role it played? 3 6. In the lab this term, copper sulphate solution and arsenomolybdate solution were used in the same experiment. Name the experiment and briefly explain the purpose and mode of action of these reagents.

4 April 10, 2006, 1:30 AM - 4:30 PM Page 4 (of 4) Biochemistry II Laboratory Section Final Examination Examiner: Dr. A. Scoot 5 7. Briefly describe the assay used in the lab this term for lysozyme. Your answer should indicate the reagents used and include a description of their mode of action; specific volumes and concentrations are not required..

5 Name: Student Number: April 10, 2006, 1:30 AM - 4:30 PM Page 1 (of 5) 1. Answer ALL questions in the space provided. 2. The back side of each page may be used for your answer or for preliminary work. 3. Questions to invigilators about the exam will not be answered Describe using diagrams the mechanism by which adrenalin affects the activity of protein phosphatase and, as a result, glycogen metabolism Using diagrams, outline the Meselson Stahl experiment and the conclusions that can be drawn from it about how DNA is replicated.

6 April 10, 2006, 1:30 AM - 4:30 PM Page 2 (of 5) Using diagrams describe the processes in the ribosome including all necessary protein factors by which 1. the last peptide bond of a protein is formed by the addition of aspartic acid and 2. the completed protein is released from the ribosome. How much energy in ATP equivalents is required? 5 4. Using a graph and diagrams, describe the changes in expression of the lac operon and the molecular mechanism controlling the changes when lactose replaces glucose as the carbon source for a culture of E. coli.

7 April 10, 2006, 1:30 AM - 4:30 PM Page 3 (of 5) 5 5. Using structural formulae, compound names and enzyme names, describe the reactions that occur in C4 plants that make them different from C3 plants. What is the physiological purpose of these reactions? 5 6. Using structural formulae, describe the coenzyme used by acetylcoa carboxylase and the reaction cycle in which the coenzyme participates Using structural formulae, describe the coenzyme and its role in the transamination reaction catalyzed by branched chain amino acid transaminase.

8 April 10, 2006, 1:30 AM - 4:30 PM Page 4 (of 5) Using structural formulae and names of all intermediates and the names of enzymes and coenzymes, outline the steps required to convert an excess of valine into ammonia and palmitic acid.

9 April 10, 2006, 1:30 AM - 4:30 PM Page 5 (of 5) Using the names of all intermediates and the names of enzymes and coenzymes, outline the steps by which a bacterium would convert ammonia and acetate into UMP. Provide the structures only for the five reactions that build the uracil ring. Assume an abundant supply of coenzymes and CO 2 and the availability of small amounts of TCA cycle intermediates.