Chapter Review in Chapter 22 Biosynthesis of Amino Acids, Nucleotides, and Related Molecules

Chapter Review

KEY TERMS

Terms in bold are defined in the glossary.

Problems

1. ATP Consumption by Root Nodules in Legumes Bacteria residing in the root nodules of the pea plant consume more than 20% of the ATP produced by the plant. Suggest why these bacteria consume so much ATP.
2. Nitrate Fertilizers and Oceanic Dead Zones Farmers apply industrially fixed nitrogen, in the form of ammonia or nitrate, to agricultural fields worldwide to increase crop yields. Agricultural runoff feeds into rivers and creates large hypoxic dead zones at the point where rivers meet oceans. How does an increase in soluble fixed nitrogen create dead zones?
3. PLP Reaction Mechanisms Pyridoxal phosphate (PLP) can help catalyze transformations one or two carbons removed from the α carbon of an amino acid. The enzyme threonine synthase promotes the PLP-dependent conversion of phosphohomoserine to threonine. Suggest a mechanism for this reaction.
4. Transformation of Aspartate to Asparagine There are two routes for transforming aspartate to asparagine at the expense of ATP. Many bacteria have an asparagine synthetase that uses ammonium ion as the nitrogen donor. Mammals have an asparagine synthetase that uses glutamine as the nitrogen donor. Given that the latter requires an extra ATP (for the synthesis of glutamine), why do mammals use this route?
5. Equation for the Synthesis of Aspartate from Glucose Write the net equation for the synthesis of aspartate (a nonessential amino acid) from glucose, carbon dioxide, and ammonia.
6. Asparagine Synthetase Inhibitors in Leukemia Therapy Mammalian asparagine synthetase is a glutamine-dependent amidotransferase. Efforts to identify an effective inhibitor of human asparagine synthetase for use in chemotherapy for patients with leukemia have focused not on the amino-terminal glutaminase domain but on the carboxyl-terminal synthetase active site. Explain why the glutaminase domain is not a promising target for a useful drug.
7. Phenylalanine Hydroxylase Deficiency and Diet Tyrosine is normally a nonessential amino acid, but individuals with a genetic defect in phenylalanine hydroxylase require tyrosine in their diet for normal growth. Explain.
8. Arginine Biosynthesis The first step of arginine biosynthesis from glutamate acetylates glutamate on the α-amino group. A subsequent step late in the same pathway removes the added acetyl group. What chemical problem is solved by adding and then removing an acetyl group, with none of the acetyl atoms appearing in the arginine product of the pathway?
9. Cofactors for One-Carbon Transfer Reactions Most one-carbon transfers are promoted by one of three cofactors: biotin, tetrahydrofolate, or S-adenosylmethionine. S-Adenosylmethionine generally serves as a methyl group donor; the transfer potential of the methyl group in $N^{5}$ $upper N Superscript 5$ -methyltetrahydrofolate is insufficient for most biosynthetic reactions. However, one example of the use of $N^{5}$ $upper N Superscript 5$ -methyltetrahydrofolate in methyl group transfer is in methionine formation by the methionine synthase reaction; methionine is the immediate precursor of S-adenosylmethionine (see Fig. 18-18).

Explain how the methyl group of S-adenosylmethionine can be derived from $N^{5}$ $upper N Superscript 5$ -methyltetrahydrofolate, even though the transfer potential of the methyl group in $N^{5}$ $upper N Superscript 5$ -methyltetrahydrofolate is one-thousandth of that of S-adenosylmethionine.
10. Concerted Regulation in Amino Acid Biosynthesis Various products of glutamine metabolism independently modulate the glutamine synthetase of E. coli (see Fig. 22-8). In this concerted inhibition, the extent of enzyme inhibition is greater than the sum of the separate inhibitions caused by each product. For E. coli grown in a medium rich in histidine, what is the advantage of concerted inhibition?
11. Relationship between Folic Acid Deficiency and Anemia Folic acid deficiency, believed to be the most common vitamin deficiency, causes a type of anemia in which hemoglobin synthesis is impaired and erythrocytes do not mature properly. What is the metabolic relationship between hemoglobin synthesis and folic acid deficiency?
12. Synthesis of Polyamines The metabolic amino acid ornithine is a direct precursor of the polyamine putrescine, shown here.

$H_{3} \overset{+}{N} — {CH}_{2} — {CH}_{2} — {CH}_{2} — {CH}_{2} — \overset{+}{N} H_{3}$ $upper H Subscript 3 Baseline ModifyingAbove upper N With plus em-dash CH Subscript 2 Baseline em-dash CH Subscript 2 Baseline em-dash CH Subscript 2 Baseline em-dash CH Subscript 2 Baseline em-dash ModifyingAbove upper N With plus upper H Subscript 3 Baseline$

Subsequent reactions convert putrescine to spermine and spermidine. What type of reaction is required to convert ornithine to putrescine, and what enzymatic cofactor is needed?
13. Nucleotide Biosynthesis in Amino Acid Auxotrophic Bacteria Wild-type E. coli cells can synthesize all 20 common amino acids, but some mutants, called amino acid auxotrophs, are unable to synthesize a specific amino acid and require its addition to the culture medium for optimal growth. Besides their role in protein synthesis, some amino acids are also precursors for other nitrogenous cell products. Consider the three amino acid auxotrophs that are unable to synthesize glycine, glutamine, and aspartate, respectively. For each mutant, what nitrogenous products other than proteins would the cell fail to synthesize?
14. Inhibitors of Nucleotide Biosynthesis Suggest a mechanism for the inhibition of alanine racemase by l-fluoroalanine.
15. Mode of Action of Sulfa Drugs Some bacteria require p-aminobenzoate in the culture medium for normal growth, and their growth is severely inhibited by the addition of sulfanilamide, one of the earliest sulfa drugs. Moreover, in the presence of this drug, 5-aminoimidazole-4-carboxamide ribonucleotide (AICAR; see Fig. 22-35) accumulates in the culture medium. Addition of excess p-aminobenzoate reverses these effects.
1. What is the role of p-aminobenzoate in these bacteria? (Hint: See Fig. 18-16.)
2. Why does AICAR accumulate in the presence of sulfanilamide?
3. Why does addition of excess p-aminobenzoate reverse the inhibition and accumulation?
16. Purine Biosynthesis Which atoms of the purine ring derive from the amide nitrogen of glutamine?
1. N-1
2. N-3
3. N-7
4. N-9
17. Pathway of Carbon in Pyrimidine Biosynthesis Predict the locations of $^{14} C$ $Superscript 14 Baseline zero width space upper C$ in orotate isolated from cells grown on a small amount of uniformly labeled $[^{14} C]$ $left-bracket Superscript 14 Baseline zero width space upper C right-bracket$ succinate. Justify your prediction.
18. Nucleotides as Poor Sources of Energy Under starvation conditions, organisms can use proteins and amino acids as sources of energy. Deamination of amino acids produces carbon skeletons that can enter the glycolytic pathway and the citric acid cycle to produce energy in the form of ATP. Nucleotides are not similarly degraded for use as energy-yielding fuels. What observations about cellular physiology support this statement? What aspect of the structure of nucleotides makes them a relatively poor source of energy?
19. Treatment of Gout Physicians use allopurinol (see Fig. 22-50), an inhibitor of xanthine oxidase, to treat chronic gout. Explain the biochemical basis for this treatment. Patients treated with allopurinol sometimes develop xanthine stones in the kidneys, although the incidence of kidney damage is much lower than in untreated gout. Explain this observation in light of these solubilities in urine: uric acid, 0.15 g/L; xanthine, 0.05 g/L; and hypoxanthine, 1.4 g/L.
20. Antibiotics That Inhibit Dihydrofolate Reductase Trimethoprim, a commonly used antibiotic, inhibits the bacterial form of dihydrofolate reductase much more than it inhibits the mammalian enzyme. What metabolic processes described in this chapter are affected by depleting tetrahydrofolate?

DATA ANALYSIS PROBLEM

21. Use of Modern Molecular Techniques to Determine the Synthetic Pathway of a Novel Amino Acid Most of the biosynthetic pathways described in this chapter were determined before the development of recombinant DNA technology and genomics, so the techniques were quite different from those that researchers would use today. Here we explore an example of the use of modern molecular techniques to investigate the pathway of synthesis of a novel amino acid, (2S)-4-amino-2-hydroxybutyrate (AHBA). The techniques mentioned here are described in various places in the text; this problem is designed to show how they can be integrated in a comprehensive study.

AHBA is a γ-amino acid that is a component of some aminoglycoside antibiotics, including the antibiotic butirosin. Antibiotics modified by the addition of an AHBA residue are often more resistant to inactivation by bacterial antibiotic-resistance enzymes. As a result, understanding how AHBA is synthesized and added to antibiotics is useful in the design of pharmaceuticals.

In an article published in 2005, Li and coworkers describe how they determined the synthetic pathway of AHBA from glutamate.
1. Briefly describe the chemical transformations needed to convert glutamate to AHBA. At this point, don’t be concerned about the order of the reactions.
  Li and colleagues began by cloning the butirosin biosynthetic gene cluster from the bacterium Bacillus circulans, which makes large quantities of butirosin. They identified five genes that are essential for the pathway: btrI, btrJ, btrK, btrO, and btrV. They cloned these genes into E. coli plasmids that allow overexpression of the genes, producing proteins with “histidine tags” fused to their amino termini to facilitate purification (see p. 313).
  
  The predicted amino acid sequence of the BtrI protein showed strong homology to known acyl carrier proteins (see Fig. 21-5). Using mass spectrometry, Li and colleagues found a molecular mass of 11,812 for the purified BtrI protein (including the His tag). When the purified BtrI was incubated with coenzyme A and an enzyme known to attach CoA to other acyl carrier proteins, the majority molecular species had an $M_{r}$ $upper M Subscript r$ of 12,153.

References

Li, Y., N.M. Llewellyn, R. Giri, F. Huang, and J.B. Spencer. 2005. Biosynthesis of the unique amino acid side chain of butirosin: possible protective-group chemistry in an acyl carrier protein–mediated pathway. Chem. Biol. 12:665–675.