Tuesday, 26 May 1998, 7:00 - 10:00 p. m., 208 Gore Hall

H. B. White - Instructor

Important - Please read this before you turn the page

• Write your name on each page.
• You have a maximum of 2 hours for this part of the examination.
• Part I of the examination has the following format and point distribution.

B. (44 points) Short Answer Questions

C. (10 points) Essay Question

D. (10 points) Problem to Solve

Part II has one problem worth 25 points.

• You may refer to your notes, course reader, handouts, textbook, and any graded homework assignments. The classroom library may be used but only briefly to lookup specific information with my permission. Books from the University library are not permitted.
• If you complete Part I early, you may take a break and return at 9:00 when Part II begins.
• Part I of this examination attempts to assess your learning, problem-solving skills, and ability to communicate clearly. Writing reflects how you think. Among the "right answers" I will read, some will be better than others because they show a greater depth of understanding, provide a more logical structure, and use words with precision. Better quality answers will receive higher marks. Therefore, organize your thoughts before you write.


Graded examinations may be picked up Friday morning, 29 May.


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_______________________ 1. The organ in your body that produces trypsin.

_______________________ 2. The phylum or kingdom to which Plasmodium falciparum belongs.

_______________________ 3. Amino acid that is a biosynthetic precursor of quinine.

_______________________ 4. Disease closely related to syphilis.

_______________________ 5. Other than red blood cells, the organ or tissue in humans that Plasmodium infects.

_______________________ 6. Isoelectric point of peptide 4 from HbS relative to peptide 4 from HbA.

_______________________ 7. Transcription forms and translation decodes this class of molecules?

_______________________ 8. A human genetic disease, other than sickle cell anemia, that affects hemoglobin.

_______________________ 9. Two radioisotopes used in biochemistry. (Need two for credit)

_______________________ 10. Name for S₂O₄⁼.

_______________________ 11. Average life span of a human red blood cell.

B. (Short narrative response questions, 6 or 8 points apiece)

1. In the classroom demonstration on Friday 8 May, with the room darkened, I shone a UV light on diluted samples of egg white from two strains of chicken and successively added ~1ml portions of a dilute riboflavin solution.

a. (3 points) Describe what was observed and how the observations differed for the two egg white solutions.

b. (3 points) What is the biochemical interpretation of the observations?

2. (6 points) As described in class, Shemin and Rittenberg (1946) observed unexpectedly that nitrogen from glycine (2-amino acetic acid) gets incorporated into the four nitrogen atoms of the heme group in hemoglobin. Subsequent studies into the biosynthesis of heme revealed that the methylene carbon of glycine gets incorporated into heme but the carboxyl group does not. Furthermore, and unexpectedly, eight, rather than four, carbon atoms in heme were labeled by methylene carbon atoms from glycine. How might these results for both carbon atoms of glycine be interpreted in relation to the results for nitrogen? (Full credit for any sensible clearly explained answer. Diagrams encouraged.)

3. (6 points) Dr. Ronald Breslow, voted one of the 75 most influential chemists of the 20^th century by C & E News, spoke at the University of Delaware recently. In his talk, he described some studies with MEL (mouse erythroleukemia) cells, the same cells used in the Elnitski et al.1997 article. These cells are infected with the MEL virus that causes the cells to be come malignant and prevents them from differentiating into normal red blood cells. Following up on an observation by others, Breslow and coworkers have synthesized several compounds that, at nanomolar concentrations, cause MEL cells to differentiate into red blood cells and produce hemoglobin. These same compounds cause other types of tumor cells to cease proliferating. If you were employed at the National Cancer Institute and were asked to follow up on these observations, what would you want to know or do? Let you curiosity show. Make a list of 6 well focused, substantive questions (learning issues) relating to information in this paragraph.

4. (6 points) Allison used metabisulfite, Pauling et al. used dithionite, and Diggs et al. and Stokes just waited to accomplish the same thing. What was it that they accomplished each in their own way sometimes for different reasons?

5. (6 points) Fluoro 2,4-dinitrobenzene (Sanger's Reagent) reacts with Ingram's peptide 4 in two places. Without drawing the whole peptide, give a chemical representation of the reaction product.

6. (6 points) Given that Dr. Hardison and coworkers study the transriptional regulation of hemoglobin gene expression. Answer one of the following: Why is it that they substituted a luciferase gene in place of the hemoglobin gene on the plasmid they used in transfection experiments? Or, Explain the basis for homology searches to identify regulatory sites.

7. (8 points) Figure 1 in Ingram's first paper shows the time course for trypsin digestion of normal and sickle cell hemoglobins. He monitored the reaction by measuring the moles of NaOH per mole of hemoglobin required to maintain the pH of the reaction mixture at 8 in a pH-stat. With structures and words, explain how the production of protons is related to hemoglobin digestion by trypsin.

Bonus question: (4 points) What relation, if any, is there between the moles of protons produced and the moles of new peptides formed by trypsin digestion?

C. Essay question (10 points)

Sydney Brenner once wrote, "Progress in science depends on new techniques, new discoveries, and new ideas, probably not in that order." Consider the articles you have read for this class and reflect on Brenner's comments. Write a short coherent essay referring to two or three of those articles and relating the scientific progress reported in each to the importance of new techniques, new discoveries, and/or new ideas. Please collect your thoughts before you write. Use the back of this page if you need more space.

D. (10 points total) Dynamic equilibria and hemoglobin electrophoresis.

1. (4 points) Hemoglobin in the red blood cells of a person with sickle cell trait is now known to exist in a dynamic equilibrium between the tetrameric forms [a₂B₂ (HbA) , a₂BB^S, and a₂B^S₂ (HbS)] and small amounts of the heterodimers. Provide a diagram (no words, please) that clearly represents the dynamic equilibrium described.

2. (2 points) If the relative amounts of the tetrameric species of hemoglobin approximate a binomial distribution, what is the approximate equilibrium proportion of a₂BB^S in red blood cells from a person with sickle cell trait? Show basis for estimate.

3. (4 points) Pauling et al. (1949) and we in class observed two, not three, electrophoretic forms of hemoglobin in the blood from people with sickle cell trait - HbA and HbS. The two forms accounted for all of the hemoglobin present. Provide a reasonable explanation for the absence of the hybrid tetramer (a₂BB^S) halfway between HbA and HbS on electrophoresis.

Tuesday 26 May 1998, 9:00 - 10:00 PM

H. B. White - Instructor

• For the group part of this examination, you will reanswer the last question from Part I

• This part of the examination is worth a total of 25 points.
• Your group has 60 minutes to complete this part of the examination.
• As was the case for Part I, you may refer to your notes, course reader, handouts, and any graded homework assignment. Textbooks and the classroom library may be used. Library books are not permitted.

• The objective of Part II is to come to consensus within your group. However, if you find that you do not agree with your colleagues, you may submit a separate answer that will be graded and used in place of the group grade.

• Good Luck! Have an enjoyable and safe summer.

(25 points total) Dynamic equilibria and hemoglobin electrophoresis.

1. (10 points) Hemoglobin in the red blood cells of a person with sickle cell trait is now known to exist in a dynamic equilibrium between the tetrameric forms [a₂B₂ (HbA) , a₂BB^S, and a₂B^S₂ (HbS)] and small amounts of the heterodimers. Provide a diagram (no words, please) that clearly represents the dynamic equilibrium described.

2. (5 points) If the relative amounts of the tetrameric species of hemoglobin approximate a binomial distribution, what is the approximate equilibrium proportion of a₂BB^S in red blood cells from a person with sickle cell trait? Show basis for estimate.

3. (10 points) Pauling et al. (1949) and we in class observed two, not three, electrophoretic forms of hemoglobin in the blood from people with sickle cell trait - HbA and HbS. The two forms accounted for all of the hemoglobin present. Provide a reasonable explanation for the absence of the hybrid tetramer (a₂BB^S) halfway between HbA and HbS on electrophoresis.