Aromatic Amino Acid Metabolism in Bacillus subtilis

Paul Gollnick; Paul Babitzke; Enrique Merino; Charles Yanofsky

doi:10.1128/9781555817992.ch17

Chapter 17 : Aromatic Amino Acid Metabolism in Bacillus subtilis

Authors: Paul Gollnick, Paul Babitzke, Enrique Merino, Charles Yanofsky

Content Type: Monograph

Publication Year: 2002

Category: Bacterial Pathogenesis

Book DOI: 10.1128/9781555817992

Chapter DOI: 10.1128/9781555817992.ch17

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Abstract:

Most proteins of Bacillus subtilis contain the three aromatic amino acids--phenylalanine, tyrosine, and tryptophan--all of which are synthesized from a common precursor, chorismate. The known and putative B. subtilis genes of aromatic amino acid metabolism, their location and presumed organization in operons, and their likely enzyme products or function are summarized in this chapter. Important distinctions between B. subtilis and Escherichia coli with regard to aromatic amino acid metabolism are the existence of a multifunctional aromatic supraoperon in B. subtilis and this organism's use of cross-pathway regulation of gene expression. Trp RNA-binding attenuation protein (TRAP) activation is reduced when the hydrogen bond between Tht52 and the catboxyl group of tryptophan is prevented. The affinity of TRAP for tryptophan increases in the presence of bound RNA. In B. subtilis, the biosynthetic pathways for folate and tryptophan are highly interconnected. First, both pathways use chorismate and glutamine to synthesize the p-aminobenzoic acid and o-aminobenzoic acid precursors of folate and tryptophan, respectively. Next, the TrpG polypeptide functions as the glutamine amidotransferase component of two enzyme complexes. Further, the mtrAB operon encodes TRAP (mtrB) and GTP cyclohydrolase I (mrrA). Finally, TRAP negatively regulates both biosynthetic pathways in response to tryptophan. The chapter talks about tRNA trp regulation of trp gene expression, a gene coding for a putative tryptophan transport protein, and homologous polypeptides in some other bacterial species.

Citation: Gollnick P, Babitzke P, Merino E, Yanofsky C. 2002. Aromatic Amino Acid Metabolism in Bacillus subtilis, p 233-244. In Sonenshein A, Losick R, Hoch J (ed), Bacillus subtilis and Its Closest Relatives. ASM Press, Washington, DC. doi: 10.1128/9781555817992.ch17

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Aromatic Amino Acid Metabolism in Bacillus subtilis

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FIGURE 1

Enzymes, genes, and reactions of the common aromatic pathway leading to the synthesis of chorismate, the precursor of phenylalanine, tyrosine, tryptophan, and several other aromatic compounds.

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FIGURE 1

Enzymes, genes, and reactions of the common aromatic pathway leading to the synthesis of chorismate, the precursor of phenylalanine, tyrosine, tryptophan, and several other aromatic compounds.

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FIGURE 2

Enzymes, genes, and reactions of the pathways leading to the synthesis of phenylalanine and tyrosine from chorismate.

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10.1128/9781555817992/fig17-2.gif

FIGURE 2

Enzymes, genes, and reactions of the pathways leading to the synthesis of phenylalanine and tyrosine from chorismate.

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FIGURE 3

Enzymes, genes, and reactions of the pathway leading to the synthesis of tryptophan from chorismate.

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FIGURE 3

Enzymes, genes, and reactions of the pathway leading to the synthesis of tryptophan from chorismate.

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FIGURE 4

Transcription attenuation model for the trpEDCFBA operon.

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FIGURE 4

Transcription attenuation model for the trpEDCFBA operon.

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Figure 5

Translational control model for trpE.

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Figure 5

Translational control model for trpE.

References

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4. Babitzke, P.,, and C. Yanofsky. 1993. Reconstitution of Bacillus subtilis trp attenuation in vitro with TRAP, the trp RNA-binding attenuation protein. Proc. Natl. Acad. Sci. USA 90: 133– 137.

5. Babitzke, P.,, and C. Yanofsky. 1995. Structural features of L-tryptophan required for activation of TRAP, the trp RNA-binding attenuation protein of Bacillus subtilis. J. Biol. Chem. 270: 12452– 12456.

6. Babitzke, P.,, D. G. Bear,, and C. Yanofsky. 1995. TRAP, the trp RNA-binding attenuation protein of Bacillus subtilis, is a toroid-shaped molecule that binds transcripts containing GAG or UAG repeats separated by two nucleotides. Proc. Nad. Acad. Sci. USA 92: 7916– 7920.

7. Babitzke, P.,, J. T. Stults,, S. J. Shire,, and C. Yanofsky. 1994. TRAP, the trp RNA-binding attenuation protein of Bacillus subtilis, is a multisubunit complex that appears to recognize G/UAG repeats in the trpEDCFBA and trpG transcripts. J. Biol. Chem. 269: 16597– 16604.

8. Babitzke, P.,, J. Yealy,, and D. Campanelli. 1996. Interaction of the trp RNA-binding attenuation protein (TRAP) of Bacillus subtilis with RNA: effects of the number of GAG repeats, the nucleotides separating adjacent repeats, and RNA secondary structure. J. Bacteriol. 178: 5159– 5163.

9. Babitzke, P.,, P. Gollnick,, and C. Yanofsky. 1992. The mtrAB operon of Bacillus subtilis encodes GTP cyclohydrolase I (MtrA), an enzyme involved in folic acid biosynthesis, and MtrB, a regulator of L-tryptophan biosynthesis. J. Bacteriol. 174: 2059– 2064.

10. Baumann, C.,, J. Otridge,, and P. Gollnick. 1996. Kinetic and thermodynamic analysis of the interaction between TRAP (trp RNA-binding attenuation protein) of Bacillus subtilis and trp leader RNA. J. Biol. Chem. 271: 12269– 12274.

11. Baumann, C.,, S. Xiragasar,, and P. Gollnick. 1997. The trp RNA-binding attenuation protein (TRAP) from Bacillus subtilis binds to unstacked trp leader RNA. J. Biol. Chem. 272: 19863– 19869.

12. Chen, X.-P.,, A. A. Antson,, M. Yang,, P. Li,, C. Baumann,, E. J. Dodson,, G. G. Dodson,, and P. Gollnick. 1999. Regulatory features of the trp operon and the crystal structure of the trp RNA-binding attenuation protein from Bacillus stearothermophilus. J. Mol. Biol. 289: 1003– 1016.

13. de Saizieu, A.,, P. Vankan,, C. Vockler,, and A. P. G. M. van Loon. 1997. The trp RNA-binding attenuation protein (TRAP) regulates the steady-state levels of transcripts of the Bacillus subtilis folate operon. Microbiology 143: 979– 989.

14. Du, H.,, and P. Babitzke. 1998. trp-RNA binding attenuation protein-mediated long-distance RNA refolding regulates translation of trpE in Bacillus subtilis. J. Biol. Chem. 273: 20494– 20503.

15. Du, H.,, A. V. Yakhnin,, S. Dharmaraj,, and P. Babitzke. 2000. trp RNA-binding attenuation protein-5′ stem-loop RNA interaction is required for proper transcription attenuation control of the Bacillus subtilis trpEDCFBA operon. J. Bacteriol 182: 1819– 1827.

16. Du, H.,, R. Tarpey,, and P. Babitzke. 1997. The trp-RNA binding attenuation protein regulates TrpG synthesis by binding to the trpG ribosome binding site of Bacillus subtilis. J. Bacteriol. 179: 2582– 2586.

17. Elliott, M. B.,, P. A. Gottlieb,, and P. Gollnick. 1999. Probing the TRAP-RNA interaction with nucleoside analogs. RNA 5: 1277– 1289.

18. Fajardo-Cavazos, P.,, and W. L. Nicholson. 2000. The TRAP-like SplA protein is a trans-acting negative regulator of spore photoproduct lyase synthesis during Bacillus subtilis sporulation. J. Bacteriol. 182: 555– 560.

19. Gollnick, P. 1994. Regulation of the Bacillus subtilis trp operon by an RNA-binding protein. Mol. Microbiol. 11: 991– 997.

20. Gollnick, P.,, S. Ishino,, M. I. Kuroda,, D. J. Henner,, and C. Yanofsky. 1990. The mtr locus is a two gene operon required for transcription attenuation in the trp operon of Bacillus subtilis. Proc. Nad. Acad. Sci. USA 87: 8726– 8730.

21. Harvey, R. J. 1973. Growth of initiation of protein synthesis in Escherichia coli in the presence of trimethoprim. J. Bacteriol. 114: 309– 322.

22. Henkin, T. M. 1996. Control of transcription termination in prokaryotes. Annu. Rev. Genet. 30: 35– 57.

23. Henner, D.,, and C. Yanofsky,. 1993. Biosynthesis of aromatic amino acids, p. 269– 280. In A. L. Sonenshein,, J. A. Hoch,, and R. Losick (ed.), Bacillus subtilis and Other Gram-Positive Bacteria: Biochemistry, Physiohgy and Molecular Genetics. American Society for Microbiology, Washington, D.C.

24. Henner, D.,, L. Band,, and H. Shimotsu. 1984. Nucleotide sequence of the Bacillus subtilis tryptophan operon. Gene 34: 169– 177.

25. Hoffman, R. J.,, and P. Gollnick. 1995. The mtrB gene of Bacillus pumilus encodes a protein with sequence and functional homology to the trp RNA-binding attenuation protein (TRAP) of Bacillus subtilis. J. Bacteriol. 177: 839– 842.

26. Kane, J. F. 1977. Regulation of a common amidotrans-ferase subunit. J. Bacteriol. 132: 419– 425.

27. Kuroda, M. I.,, D. Henner,, and C. Yanofsky. 1988. cis-acting sites in the transcript of the Bacilus subtilis trp operon regulate expression of the operon. J. Bacteriol. 170: 3080– 3088.

28. Landick, R.,, C. L. Turnbough, Jr.,, and C. Yanofsky,. 1996. Transcription attenuation, p. 1263– 1286. In F. C. Neidhardt,, R. Curtiss III,, J. L. Ingraham,, E. C. C. Lin,, K. B. Low, Jr.,, B. Maganasik,, W. S. Reznikoff,, M. Riley,, M. Schaechter,, and H. E. Umbarger (ed.), Escherichia coli and Salmonella: Cellular and Molecuhr Biology. American Society for Microbiology, Washington, D.C.

29. Lee, A. I.,, J. P. Sarsero,, and C. Yanofsky. 1996. A temperature-sensitive trpS mutation interferes with TRAP regulation of trp gene expression in Bacillus subtilis. J. Bacteriol. 178: 6518– 6524.

30. Merino, E.,, P. Babitzke,, and C. Yanofsky. 1995. trp RNA-binding attenuation protein (TRAP)- trp leader RNA interactions mediate translational as well as transcriptional regulation of the Bacillus subtilis trp operon. J. Bacteriol. 177: 6362– 6370.

31. Otridge, J.,, and P. Gollnick. 1993. MtrB from Bacillus subtilis binds specifically to trp leader RNA in a tryptophan dependent manner. Proc. Natl. Acad. Sci. USA 90: 128– 132.

32. Pearson, W. R. 2000. Flexible sequence similarity searching with the FASTA3 program package. Methods Mol. Biol. 132: 185– 219.

33. Sarsero, J. P.,, E. Merino,, and C. Yanofsky. 2000. A Bacillus subtilis gene of previously unknown function, yhaG, is translationally regulated by tryptophan-activated TRAP and appears to be involved in tryptophan transport. J. Bacteriol. 182: 2329– 2331.

34. Sarsero, J.P.,, E. MerinoandC.Yanofsky. 2000. A Bacillus subtilis operon containing genes of unknown function senses tRNA Trp charging and regulates expression of the genes of tryptophan biosynthesis. Proc. Natl. Acad. Sci. USA 97: 2656– 2661.

35. Shimotsu, H.,, and D. J. Henner. 1984. Characterization of the Bacillus subtilis tryptophan promoter region. Proc. Natl. Acad. Sci. USA 43: 85– 94.

36. Shimotsu, H.,, and D. J. Henner. 1986. Construction of a single-copy integration vector and its use in analysis of regulation of the trp operon of Bacillus subtilis. Gene 43: 85– 94·

37. Shimotsu, H.,, M. I. Kuroda,, C. Yanofsky,, and D. J. Henner. 1986. Novel form of transcription attenuation regulates expression of the Bacillus subtilis tryptophan operon. J. Bacteriol. 166: 461– 471.

38. Slock, J.,, D. P. Stahly,, C.-Y. Han,, E. W. Six,, and I. P. Crawford. 1990. An apparent Bacillus subtilis folic acid biosynthetic operon containing pab, an amphibolic trpG gene, a third gene required for synthesis of para-aminobenzoic acid, and the dihydropteroate synthase gene. J. Bacteriol. 172: 7211– 7226.

39. Steinberg, W. 1974. Temperature-induced derepression of tryptophan biosynthesis in a tryptophanyl-transfer ribonucleic acid synthetase mutant of Bacillus subtilis. J. Bacteriol. 117: 1023– 1034.

40. Sudershana, S.,, H. Du,, M. Mahalanabis,, and P. Babitzke. 1999. A 5′ RNA stem-loop participates in the transcription attenuation mechanism that controls expression of the Bacillus subtilis trpEDCFBAopeton. J. Bacteriol. 181: 5742– 5749.

41. Xirasager, S.,, M. B. Elliott,, W. Bartolini,, P. Gollnick,, and P. Gottlieb. 1998. RNA structure inhibits the TRAP ( trp RNA-binding attenuation protein)-RNA interaction. J. Biol. Chem. 273: 27146– 27153.

42. Yakhnin, A. V.,, J. J. Trimble,, C. R. Chiaro,, and P. Babitzke. 2000. Effects of mutations in the L-tryptophan binding pocket of the trp RNA-binding attenuation protein of Bacillus subtilis. J. Biol. Chem. 275: 4519– 4524.

43. Yakhnin, H. B.,, A. V. Yakhnin,, and P. Babitzke. Unpublished data.

44. Yang, M.,, A. de Saizieu,, A. P. G. M. van Loon,, and P. Gollnick. 1995. Translation of trpG in Bacillus subtilis is regulated by the trp RNA-binding attenuation protein (TRAP). J. Bacteriol. 177: 4272– 4278.

45. Yang, M.,, X.-P. Chen,, and P. Gollnick. 1997. Alanine-scanning mutagenesis of Bacillus subtilis trp-RNA binding attenuation protein (TRAP) reveals residues involved in tryptophan binding and RNA binding. J. Mol. Biol. 270: 696– 710.

Tables

Aromatic Amino Acid Metabolism in Bacillus subtilis

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TABLE 1

Operon organization of the genes of aromatic amino acid metabolism a

a Genes for tRNAs and tRNA synthecases are not shown.

b Map position reflects the relative gene position on the chromosome map.

c Transcription orientation. F, forward, clockwise; R, reverse, counterclockwise.

d Intergenic spacing (negative number indicates genes overlap); or number of nucleotides from the promoter to the first gene of the operon.

e Alternative name: para-aminobenzoate synthase glutamine amidotransferase subunit B.

f Transcription also starts from the trpE promoter localized 203 bp upstream from its ATG start codon.

g Alternative name: L-serine hydrolyase.

h Alternative name: indole-3-glycerol phosphate aldolase.

i Alternative name: hisC.

j Alternative name: 3-phosphoshikimate 1-carboxyvinyltransferase.

k Alternative name: aroG. Bifunctional enzyme. DAHP syntase/chorismate mutase.

l Alternative name: phospho-2-dehydro-3'deoxyheptonate aldolase.

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TABLE 1

Operon organization of the genes of aromatic amino acid metabolism a

a Genes for tRNAs and tRNA synthecases are not shown.

b Map position reflects the relative gene position on the chromosome map.

c Transcription orientation. F, forward, clockwise; R, reverse, counterclockwise.

d Intergenic spacing (negative number indicates genes overlap); or number of nucleotides from the promoter to the first gene of the operon.

e Alternative name: para-aminobenzoate synthase glutamine amidotransferase subunit B.

f Transcription also starts from the trpE promoter localized 203 bp upstream from its ATG start codon.

g Alternative name: L-serine hydrolyase.

h Alternative name: indole-3-glycerol phosphate aldolase.

i Alternative name: hisC.

j Alternative name: 3-phosphoshikimate 1-carboxyvinyltransferase.

k Alternative name: aroG. Bifunctional enzyme. DAHP syntase/chorismate mutase.

l Alternative name: phospho-2-dehydro-3'deoxyheptonate aldolase.

/content/10.1128/9781555817992.chap17.tab17-2

TABLE 2

Polypeptide homologs in some other species (percent identity of the amino acid sequence with the B. subtilis sequence) a

a The fully sequenced genomes analyzed were B. subtilis (AL009126), E. coli (U00096), Deinococcus radiodurcms chromosomes 1 (AE000513) and 2 (AE001825), Methanobacterium thermoautotrophicum (AE000666), Mycoplasma genitalium (L43967), Mycoplasma pneumoniae (U00089), and Mycobacterium tuberculosis (AL123456). Partially sequenced genomes are from NCBI-GenBank Flat File Release 109.0. Owing to the multidomain nature of some enzymes in the pathways, comparisons were restricted to common functional domains using the local alignment program ssearch3, designed for full-length protein comparisons ( 32 ). No homologs of B. subtilis YhaG, YczA, and YcbK were found; therefore, the corresponding proteins were omitted from this table.

b Bifunctional enzyme: DAHP syntase/chorismate mutase.

c N.s., there is biochemical evidence of this activity, but the search did not find a statistically significant similarity.

d There are three different versions of this enzyme.

e N.f., not found.

f Bifunctional enzyme: chorismate mutase/prephenate dehydratase.

g Bifunctional enzyme: chorismate mutase/prephenate dehydrogenase.

h This pathway also includes chorismate mutase and tyrosine/phenylalanine aminotransferase.

i Bifunctional enzyme: phosphoribosyl anthranilate isomerase/indole-3'glycerol phosphate synthase.

j Bifunctional enzyme: anthranilate synthase component II/anthranilate phosphoribosyltransferase.

k Bifunctional enzyme: phosphoribosyl anthranilate isomerase/indole-3-glycerol phosphate synthase.

l mtrB has also been sequenced from Bacillus pumilus ( 25 ).

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TABLE 2

Polypeptide homologs in some other species (percent identity of the amino acid sequence with the B. subtilis sequence) a

b Bifunctional enzyme: DAHP syntase/chorismate mutase.

c N.s., there is biochemical evidence of this activity, but the search did not find a statistically significant similarity.

d There are three different versions of this enzyme.

e N.f., not found.

f Bifunctional enzyme: chorismate mutase/prephenate dehydratase.

g Bifunctional enzyme: chorismate mutase/prephenate dehydrogenase.

h This pathway also includes chorismate mutase and tyrosine/phenylalanine aminotransferase.

i Bifunctional enzyme: phosphoribosyl anthranilate isomerase/indole-3'glycerol phosphate synthase.

j Bifunctional enzyme: anthranilate synthase component II/anthranilate phosphoribosyltransferase.

k Bifunctional enzyme: phosphoribosyl anthranilate isomerase/indole-3-glycerol phosphate synthase.

l mtrB has also been sequenced from Bacillus pumilus ( 25 ).