From the beginning or extract and save and reuse. Differentiate between the de novo synthesis of nucleotides and salvage-pathway synthesis.

Finding their roots 1. Identify the source of the atoms in the pyrimidine ring

Finding their roots 2. Identify the source of the atoms in the purine ring.

Multifaceted. List some of the biochemical roles played by nucleotides.

An s instead of a t? Differentiate between a nucleoside and a nucleotide.

Associate ’em.

Safe passage. What is substrate channeling? How does it affect enzyme efficiency?

Activated ribose phosphate. Write a balanced equation for the synthesis of PRPP from glucose through the oxidative branch of the pentose phosphate pathway.

Making a pyrimidine. Write a balanced equation for the synthesis of orotate from glutamine, CO₂, and aspartate.

Identifying the donor. What is the activated reactant in the biosynthesis of each of the following compounds?

Inhibiting purine biosynthesis. Amidotransferases are inhibited by the antibiotic azaserine (O-diazoacetyl-l-serine), which is an analog of glutamine.

Which intermediates in purine biosynthesis would accumulate in cells treated with azaserine?

The price of methylation. Write a balanced equation for the synthesis of TMP from dUMP that is coupled to the conversion of serine into glycine.

Sulfa action. Bacterial growth is inhibited by sulfanilamide and related sulfa drugs, and there is a concomitant accumulation of 5-aminoimidazole-4-carboxamide ribonucleotide. This inhibition is reversed by the addition of p-aminobenzoate.

Propose a mechanism for the inhibitory effect of sulfanilamide.

HAT medium. Mutant cells unable to synthesize nucleotides by salvage pathways are very useful tools in molecular and cell biology. Suppose that cell A lacks thymidine kinase, the enzyme catalyzing the phosphorylation of thymidine to thymidylate, and that cell B lacks hypoxanthine-guanine phosphoribosyl transferase.

Vitamin requirement. What is the role of folate in pyrimidine synthesis and what is the consequence of a folate deficiency during development?

Bringing equilibrium. What is the reciprocal substrate relation in the synthesis of ATP and GTP?

Find the label. Suppose that cells are grown on amino acids that have all been labeled at the α carbons with ¹³C. Identify the atoms in cytosine and guanine that will be labeled with ¹³C.

Need a map? Describe the path for the synthesis of TTP from UTP.

Adjunct therapy. Allopurinol is sometimes given to patients with acute leukemia who are being treated with anticancer drugs. Why is allopurinol used?

A hobbled enzyme. Both side-chain oxygen atoms of aspartate 27 at the active site of dihydrofolate reductase form hydrogen bonds with the pteridine ring of folates. The importance of this interaction was assessed by studying two mutants at this position, Asn 27 and Ser 27. The dissociation constant of methotrexate was 0.07 nM for the wild type, 1.9 nM for the Asn 27 mutant, and 210 nM for the Ser 27 mutant, at 25°C. Calculate the standard free energy of the binding of methotrexate by these three proteins. What is the decrease in binding energy resulting from each mutation?

Needed supplies. Why are cancer cells especially sensitive to inhibitors of TMP synthesis?

Alternative routes. Orotic aciduria results when one of the enzyme activities of UMP synthetase is absent. This syndrome is characterized by large amounts of orotic acid in the blood and urine, megaloblastic anemia (characterized by large, immature, and dysfunctional red blood cells) and retarded growth. Suggest a possible treatment for this condition.

Correcting deficiencies. Suppose that a person is found who is deficient in an enzyme required for IMP synthesis. How might this person be treated?

Labeled nitrogen. Purine biosynthesis is allowed to take place in the presence of [¹⁵N]aspartate, and the newly synthesized GTP and ATP are isolated. What positions are labeled in the two nucleotides?

On the trail of carbons. Tissue culture cells were incubated with glutamine labeled with ¹⁵N in the amide group. Subsequently, IMP was isolated and found to contain some ¹⁵N. Which atoms in IMP were labeled?

Mechanism of action. What is the biochemical basis of allopurinol treatment for gout?

Changed inhibitor. Xanthine oxidase treated with allopurinol results in the formation of a new compound that is an extremely potent inhibitor of the enzyme. Propose a structure for this compound.

Calculate the ATP footprint. How many molecules of ATP are required to synthesize one molecule of CTP from scratch?

Blockages. What intermediate in purine synthesis will accumulate if a strain of bacteria is lacking each of the following biochemicals?

The same and not the same. Write out mechanisms for the conversion of phosphoribosylamine into glycinamide ribonucleotide and of xanthylate into guanylate.

Closing the ring. Propose a mechanism for the conversion of 5-formamidoimidazole-4-carboxamide ribonucleotide into inosinate.

Different strokes. Human beings contain two different carbamoyl phosphate synthetase enzymes. One uses glutamine as a substrate, whereas the other uses ammonia. What are the functions of these two enzymes?

A generous donor. What major biosynthetic reactions utilize PRPP?

They’re everywhere! Nucleotides play a variety of roles in the cell. Give an example of a nucleotide that acts in each of the following roles or processes.

Pernicious anemia. Purine biosynthesis is impaired by vitamin B₁₂ deficiency. Why? How might fatty acid and amino acid metabolism also be affected by a vitamin B₁₂ deficiency?

Folate deficiency. Suppose someone was suffering from a folate deficiency. What cells would you think might be most affected? Symptoms may include diarrhea and anemia.

Hyperuricemia. Many patients with glucose 6-phosphatase deficiency have high serum levels of urate. Hyperuricemia can be induced in normal people by the ingestion of alcohol or by strenuous exercise. Propose a common mechanism that accounts for these findings.

Labeled carbon. Succinate uniformly labeled with ¹⁴C is added to cells actively engaged in pyrimidine biosynthesis. Propose a mechanism by which carbon atoms from succinate could be incorporated into a pyrimidine. At what positions is the pyrimidine labeled?

Something funny going on here. Cells were incubated with glucose labeled with ¹⁴C in carbon 2, shown in red in the structure below. Later, uracil was isolated and found to contain ¹⁴C in carbons 4 and 6. Account for this labeling pattern.

Side effects. Azathioprine is an immunosuppressant drug used in kidney transplants. In vivo, azathioprine is metabolized to the hypoxanthine analog 6-mercaptopurine, which is then converted into 6-mercaptopurine ribose monophosphate, the active form of the drug. 6-Mercaptopurine ribose monophosphate also inhibits de novo purine synthesis, reducing uric acid levels in the blood and urine. However, when administered to Lesch–Nyhan patients, there was no effect on uric acid levels. Explain why this is the case.

Exercising muscle. Some interesting reactions take place in muscle tissue to facilitate the generation of ATP for contraction. In muscle contraction, ATP is converted into ADP. Adenylate kinase converts two molecules of ADP into a molecule of ATP and AMP.

A common step. What three reactions transfer an amino group from aspartate to yield the aminated product and fumarate?

Your pet duck. You suspect that your pet duck has gout. Why should you think twice before administering a dose of allopurinol-laced bread?

Not all absences are equal. In order to better understand the nature of the defects in HGPRT in patients suffering from Lesch–Nyhan disease, cell lines from two Lesch–Nyhan patients (LND 1 and LND 2) as well as a normal individual were established. HGPRT was then purified from each of the cell lines using the techniques examined in Chapter 3. Using the purified enzyme, the basic kinetics of the three enzymes were compared, as shown in Figure A. In each case, the same amount of purified enzyme was used in the analysis.