Undergraduates, do any SIX problems. Graduate students, any SEVEN.
You may use any literature source you like, and discuss the problems with each other or with me. However, your answers must be in your own words and structure drawings. I am looking for complete, well thought out answers.
1. As we described in class, dehydroquinate synthase carries out five successive reactions on DAHP: alcohol oxidation, phosphate b-elimination, carbonyl reduction, ring opening, and intramolecular aldol condensation. Write structures in organic line notation for all of the species along this pathway. Then write an organic "curly arrow" mechanism for the aldol condensation. A place to get started: Nature, 1998, 394, 299.
2. We have said that the conversion of dehydroquinate to dehydroshikimate is a cis- elimination. (a) Explain how this conclusion was obtained for the in vivo reaction; (b) in vitro, for example with POCl3 and pyridine, this would be a trans- elimination. Write a mechanism for the laboratory reaction and explain why trans- is the preferred pathway. Your sophomore organic text will help with part (b).
3. If chorismate is labeled with 14C in the =CH2 of the pyruvyl side chain, where will the label appear in phenylalanine? Show in detail the pathway to your answer; that is, write any intermediate structures, showing where the label appears.
4. What is the structure of the herbicide Roundup? How does it work? Be specific! Why is it supposedly safe to use around pets (unless you keep bacteria for pets)? Numerous crops are now engineered to be "Roundup ready", that is, resistant to Roundup. What is the alteration made in these plants? (A place to start: Healy-Fried, et al., J. Biol. Chem., 2007, 282, 32949.)
5. The molecules below all are terpenes, formed by condensations involving isopentenyl pyrophosphate. Locate the isoprene units in each.
6. We have described in class feedback inhibition, in which a compound from later in a biosynthetic sequence is an allosteric inhibitor of an earlier enzyme. What is feed-forward activation? Provide a specific example; that is a specific enzyme and a specific activator. Explain how the activation works. Then try to find the structure of the enzyme in the Protein Databank, preferably with the activator bound. Provide a Rasmol picture. (No penalty if you can't find it in the pdb.)
7. We showed the reaction of dimethylallyl pyrophosphate with isopentenyl pyrophosphate as an SN1 reaction. However, allyl substrates readily undergo SN2 reactions, much more rapidly than other 1o substrates. Why? What is the evidence that the enzymatic reaction is actually SN1? See J. Am. Chem. Soc., 1993, 115, 1245 for a start at answering this question.
8.. Fun with asterisks. If acetate is labeled with 13C in the methyl group, which carbons of squalene will contain the label? Which carbons of lanosterol? Show all of the intermediate structures that allow you to arrive at your answer.
9. Fun with curly arrows. Write the individual steps in the "miracle" shown in our scheme for the rearrangements that occur at the end of the formation of lanosterol. Hint: think pull rather than push; that is, the carbocation center "pulls" on the electrons.
10. [This is long, but not very difficult. do either part (a) or part (b)] (a) Mother Nature regulates the biosynthesis of cholesterol at the HMG-CoA reductase stage, by phosphorylation of the enzyme. It now appears that other regulatory modes may be operating as well. Read Parish, Parish, and Li, Crit. Rev. Biochem. Mol. Biol., 1999, 34, 265, and summarize its suggestions about another mode of regulation; (b) Relatively recently, squalene synthetase has become a target for development of anti-cholesterol drugs by pharmaceutical companies. Identify one such drug and explain in detail how it works. Look for a crystal structure of the enzyme with the inhibitor bound and, if one can be found, provide a picture showing the binding of the inhibitor.
11. [This also is long, but not very difficult.] Horowitz (Proc. Nat. Acad. Sci., 1945, 31, 153; Evolving Genes and Proteins, Bryson and Vogel, eds., Academic Press, 1965, p. 16) has argued that biosynthetic pathways evolved in reverse; that is, from the last step back toward the first. Summarize his arguments. Do you see any obvious defects? What are they?
12. Although it has long been assumed that isopentenyl pyrophosphate is synthesized only from mevalonate as we showed in class, another pathway recently has been discovered. The new pathway condenses glyceraldehyde 3-phosphate and pyruvate to 1-deoxyxylulose 5-phosphate. Describe the new pathway and any of the enzymes that have been identified. [A place to start: Hunter, J. Biol. Chem., 2007, 282, 21573].
13. Some effort has been expended on explaining the evolutionary relationship between the two isoprenoid pathways (problem # 12). Describe thoroughly the suggestions. [Mol. Microbiol., 2000, 37, 707; Proc. Nat. Acad. Sci., 2000, 97, 13172]
14. PGH synthase is the target enzyme of aspirin and other so-called COX2 inhibitors (NSAIDs; Vioxx is an example). What reaction is catalyzed by this enzyme? Write an organic "curly arrow" mechanism for the reaction. How does aspirin inhibit the enzyme? Outline the pathway onward from the product of this enzyme to the PGF, PGE, and PGD families of prostaglandins.