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. 1991 Apr;173(8):2590–2599. doi: 10.1128/jb.173.8.2590-2599.1991

The Bacillus subtilis hemAXCDBL gene cluster, which encodes enzymes of the biosynthetic pathway from glutamate to uroporphyrinogen III.

M Hansson 1, L Rutberg 1, I Schröder 1, L Hederstedt 1
PMCID: PMC207825  PMID: 1672867

Abstract

We have recently reported (M. Petricek, L. Rutberg, I. Schröder, and L. Hederstedt, J. Bacteriol. 172: 2250-2258, 1990) the cloning and sequence of a Bacillus subtilis chromosomal DNA fragment containing hemA proposed to encode the NAD(P)H-dependent glutamyl-tRNA reductase of the C5 pathway for 5-aminolevulinic acid (ALA) synthesis, hemX encoding a hydrophobic protein of unknown function, and hemC encoding hydroxymethylbilane synthase. In the present communication, we report the sequences and identities of three additional hem genes located immediately downstreatm of hemC, namely, hemD encoding uroporphyrinogen III synthase, hemB encoding porphobilinogen synthase, and hemL encoding glutamate-1-semialdehyde 2,1-aminotransferase. The six genes are proposed to constitute a hem operon encoding enzymes required for the synthesis of uroporphyrinogen III from glutamyl-tRNA. hemA, hemB, hemC, and hemD have all been shown to be essential for heme synthesis. However, deletion of an internal 427-bp fragment of hemL did not create a growth requirement for ALA or heme, indicating that formation of ALA from glutamate-1-semialdehyde can occur spontaneously in vivo or that this reaction may also be catalyzed by other enzymes. An analysis of B. subtilis carrying integrated plasmids or deletions-substitutions in or downstream of hemL indicates that no further genes in heme synthesis are part of the proposed hem operon.

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Selected References

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  1. Alefounder P. R., Abell C., Battersby A. R. The sequence of hemC, hemD and two additional E. coli genes. Nucleic Acids Res. 1988 Oct 25;16(20):9871–9871. doi: 10.1093/nar/16.20.9871. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Alwan A. F., Mgbeje B. I., Jordan P. M. Purification and properties of uroporphyrinogen III synthase (co-synthase) from an overproducing recombinant strain of Escherichia coli K-12. Biochem J. 1989 Dec 1;264(2):397–402. doi: 10.1042/bj2640397. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Anderson T. J., Ivánovics G. Isolation and some characteristics of haemin dependent mutants of Bacillus subtilis. J Gen Microbiol. 1967 Oct;49(1):31–40. doi: 10.1099/00221287-49-1-31. [DOI] [PubMed] [Google Scholar]
  4. Arwert F., Venema G. Transformation in Bacillus subtilis. Fate of newly introduced transforming DNA. Mol Gen Genet. 1973;123(2):185–198. doi: 10.1007/BF00267334. [DOI] [PubMed] [Google Scholar]
  5. Avissar Y. J., Beale S. I. Identification of the enzymatic basis for delta-aminolevulinic acid auxotrophy in a hemA mutant of Escherichia coli. J Bacteriol. 1989 Jun;171(6):2919–2924. doi: 10.1128/jb.171.6.2919-2924.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Bachmann B. J. Linkage map of Escherichia coli K-12, edition 8. Microbiol Rev. 1990 Jun;54(2):130–197. doi: 10.1128/mr.54.2.130-197.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Beale S. I., Castelfranco P. A. The Biosynthesis of delta-Aminolevulinic Acid in Higher Plants: II. Formation of C-delta-Aminolevulinic Acid from Labeled Precursors in Greening Plant Tissues. Plant Physiol. 1974 Feb;53(2):297–303. doi: 10.1104/pp.53.2.297. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Berek I., Miczák A., Ivánovics G. Mapping the delta-aminolaevulinic acid dehydrase and porphobilinogen deaminase loci in Bacillus subtilis. Mol Gen Genet. 1974;132(3):233–239. doi: 10.1007/BF00269396. [DOI] [PubMed] [Google Scholar]
  9. Berek I., Miczák A., Kiss I., Ivánovics G., Durkó I. Genetic and biochemical analysis of haemin dependent mutants of Bacillus subtilis. Acta Microbiol Acad Sci Hung. 1975;22(2):157–167. [PubMed] [Google Scholar]
  10. Bishop T. R., Frelin L. P., Boyer S. H. Nucleotide sequence of rat liver delta-aminolevulinic acid dehydratase cDNA. Nucleic Acids Res. 1986 Dec 22;14(24):10115–10115. doi: 10.1093/nar/14.24.10115. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Chartrand P., Tardif D., Săsărman A. Uroporphyrin- and coproporphyrin I-accumulating mutant of Escherichia coli K12. J Gen Microbiol. 1979 Jan;110(1):61–66. doi: 10.1099/00221287-110-1-61. [DOI] [PubMed] [Google Scholar]
  12. Devereux J., Haeberli P., Smithies O. A comprehensive set of sequence analysis programs for the VAX. Nucleic Acids Res. 1984 Jan 11;12(1 Pt 1):387–395. doi: 10.1093/nar/12.1part1.387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Echelard Y., Dymetryszyn J., Drolet M., Sasarman A. Nucleotide sequence of the hemB gene of Escherichia coli K12. Mol Gen Genet. 1988 Nov;214(3):503–508. doi: 10.1007/BF00330487. [DOI] [PubMed] [Google Scholar]
  14. Elliott T., Avissar Y. J., Rhie G. E., Beale S. I. Cloning and sequence of the Salmonella typhimurium hemL gene and identification of the missing enzyme in hemL mutants as glutamate-1-semialdehyde aminotransferase. J Bacteriol. 1990 Dec;172(12):7071–7084. doi: 10.1128/jb.172.12.7071-7084.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Elliott T. Cloning, genetic characterization, and nucleotide sequence of the hemA-prfA operon of Salmonella typhimurium. J Bacteriol. 1989 Jul;171(7):3948–3960. doi: 10.1128/jb.171.7.3948-3960.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Elliott T., Roth J. R. Heme-deficient mutants of Salmonella typhimurium: two genes required for ALA synthesis. Mol Gen Genet. 1989 Apr;216(2-3):303–314. doi: 10.1007/BF00334369. [DOI] [PubMed] [Google Scholar]
  17. Gibbs P. N., Jordan P. M. Identification of lysine at the active site of human 5-aminolaevulinate dehydratase. Biochem J. 1986 Jun 1;236(2):447–451. doi: 10.1042/bj2360447. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Grimm B., Bull A., Welinder K. G., Gough S. P., Kannangara C. G. Purification and partial amino acid sequence of the glutamate 1-semialdehyde aminotransferase of barley and synechococcus. Carlsberg Res Commun. 1989;54(2):67–79. doi: 10.1007/BF02907586. [DOI] [PubMed] [Google Scholar]
  19. Grimm B. Primary structure of a key enzyme in plant tetrapyrrole synthesis: glutamate 1-semialdehyde aminotransferase. Proc Natl Acad Sci U S A. 1990 Jun;87(11):4169–4173. doi: 10.1073/pnas.87.11.4169. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Hart G. J., Battersby A. R. Purification and properties of uroporphyrinogen III synthase (co-synthetase) from Euglena gracilis. Biochem J. 1985 Nov 15;232(1):151–160. doi: 10.1042/bj2320151. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Ish-Horowicz D., Burke J. F. Rapid and efficient cosmid cloning. Nucleic Acids Res. 1981 Jul 10;9(13):2989–2998. doi: 10.1093/nar/9.13.2989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Jordan P. M., Mgbeje B. I., Thomas S. D., Alwan A. F. Nucleotide sequence for the hemD gene of Escherichia coli encoding uroporphyrinogen III synthase and initial evidence for a hem operon. Biochem J. 1988 Jan 15;249(2):613–616. doi: 10.1042/bj2490613. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Kannangara C. G., Gough S. P., Bruyant P., Hoober J. K., Kahn A., von Wettstein D. tRNA(Glu) as a cofactor in delta-aminolevulinate biosynthesis: steps that regulate chlorophyll synthesis. Trends Biochem Sci. 1988 Apr;13(4):139–143. doi: 10.1016/0968-0004(88)90071-0. [DOI] [PubMed] [Google Scholar]
  24. Kieser T. Factors affecting the isolation of CCC DNA from Streptomyces lividans and Escherichia coli. Plasmid. 1984 Jul;12(1):19–36. doi: 10.1016/0147-619x(84)90063-5. [DOI] [PubMed] [Google Scholar]
  25. Kiss I., Berek I., Ivánovics G. Mapping the -aminolaevulinic acid synthetase locus in Bacillus subtilis. J Gen Microbiol. 1971 May;66(2):153–159. doi: 10.1099/00221287-66-2-153. [DOI] [PubMed] [Google Scholar]
  26. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  27. Li J. M., Russell C. S., Cosloy S. D. The structure of the Escherichia coli hemB gene. Gene. 1989 Jan 30;75(1):177–184. doi: 10.1016/0378-1119(89)90394-6. [DOI] [PubMed] [Google Scholar]
  28. Li J. M., Umanoff H., Proenca R., Russell C. S., Cosloy S. D. Cloning of the Escherichia coli K-12 hemB gene. J Bacteriol. 1988 Feb;170(2):1021–1025. doi: 10.1128/jb.170.2.1021-1025.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Liedgens W., Lütz C., Schneider H. A. Molecular properties of 5-aminolevulinic acid dehydratase from Spinacia oleracea. Eur J Biochem. 1983 Sep 1;135(1):75–79. doi: 10.1111/j.1432-1033.1983.tb07619.x. [DOI] [PubMed] [Google Scholar]
  30. Mandel M., Higa A. Calcium-dependent bacteriophage DNA infection. J Mol Biol. 1970 Oct 14;53(1):159–162. doi: 10.1016/0022-2836(70)90051-3. [DOI] [PubMed] [Google Scholar]
  31. May B. K., Borthwick I. A., Srivastava G., Pirola B. A., Elliott W. H. Control of 5-aminolevulinate synthase in animals. Curr Top Cell Regul. 1986;28:233–262. doi: 10.1016/b978-0-12-152828-7.50008-1. [DOI] [PubMed] [Google Scholar]
  32. McClung C. R., Somerville J. E., Guerinot M. L., Chelm B. K. Structure of the Bradyrhizobium japonicum gene hemA encoding 5-aminolevulinic acid synthase. Gene. 1987;54(1):133–139. doi: 10.1016/0378-1119(87)90355-6. [DOI] [PubMed] [Google Scholar]
  33. Miczák A., Prágai B., Berek I. Mapping the uroporphyrinogen III cosynthase locus in Bacillus subtilis. Mol Gen Genet. 1979 Jul 24;174(3):293–295. doi: 10.1007/BF00267802. [DOI] [PubMed] [Google Scholar]
  34. Myers A. M., Crivellone M. D., Koerner T. J., Tzagoloff A. Characterization of the yeast HEM2 gene and transcriptional regulation of COX5 and COR1 by heme. J Biol Chem. 1987 Dec 15;262(35):16822–16829. [PubMed] [Google Scholar]
  35. Neville D. M., Jr Molecular weight determination of protein-dodecyl sulfate complexes by gel electrophoresis in a discontinuous buffer system. J Biol Chem. 1971 Oct 25;246(20):6328–6334. [PubMed] [Google Scholar]
  36. Niaudet B., Goze A., Ehrlich S. D. Insertional mutagenesis in Bacillus subtilis: mechanism and use in gene cloning. Gene. 1982 Oct;19(3):277–284. doi: 10.1016/0378-1119(82)90017-8. [DOI] [PubMed] [Google Scholar]
  37. O'Neill G. P., Chen M. W., Söll D. delta-Aminolevulinic acid biosynthesis in Escherichia coli and Bacillus subtilis involves formation of glutamyl-tRNA. FEMS Microbiol Lett. 1989 Aug;51(3):255–259. doi: 10.1016/0378-1097(89)90406-0. [DOI] [PubMed] [Google Scholar]
  38. Petricek M., Rutberg L., Schröder I., Hederstedt L. Cloning and characterization of the hemA region of the Bacillus subtilis chromosome. J Bacteriol. 1990 May;172(5):2250–2258. doi: 10.1128/jb.172.5.2250-2258.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Powell K. A., Cox R., McConville M., Charles H. P. Mutations affecting porphyrin biosynthesis in Escherichia coli. Enzyme. 1973;16(1):65–73. doi: 10.1159/000459363. [DOI] [PubMed] [Google Scholar]
  40. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Sasarman A., Nepveu A., Echelard Y., Dymetryszyn J., Drolet M., Goyer C. Molecular cloning and sequencing of the hemD gene of Escherichia coli K-12 and preliminary data on the Uro operon. J Bacteriol. 1987 Sep;169(9):4257–4262. doi: 10.1128/jb.169.9.4257-4262.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Smythe E., Williams D. C. Rat liver uroporphyrinogen III synthase has similar properties to the enzyme from Euglena gracilis, including absence of a requirement for a reversibly bound cofactor for activity. Biochem J. 1988 Jul 1;253(1):275–279. doi: 10.1042/bj2530275. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Spizizen J. TRANSFORMATION OF BIOCHEMICALLY DEFICIENT STRAINS OF BACILLUS SUBTILIS BY DEOXYRIBONUCLEATE. Proc Natl Acad Sci U S A. 1958 Oct 15;44(10):1072–1078. doi: 10.1073/pnas.44.10.1072. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. Tsai S. F., Bishop D. F., Desnick R. J. Human uroporphyrinogen III synthase: molecular cloning, nucleotide sequence, and expression of a full-length cDNA. Proc Natl Acad Sci U S A. 1988 Oct;85(19):7049–7053. doi: 10.1073/pnas.85.19.7049. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. Tsukamoto I., Yoshinaga T., Sano S. The role of zinc with special reference to the essential thiol groups in delta-aminolevulinic acid dehydratase of bovine liver. Biochim Biophys Acta. 1979 Sep 12;570(1):167–178. doi: 10.1016/0005-2744(79)90211-0. [DOI] [PubMed] [Google Scholar]
  46. Wetmur J. G., Bishop D. F., Cantelmo C., Desnick R. J. Human delta-aminolevulinate dehydratase: nucleotide sequence of a full-length cDNA clone. Proc Natl Acad Sci U S A. 1986 Oct;83(20):7703–7707. doi: 10.1073/pnas.83.20.7703. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Wulff D. L. Delta-aminolevulinic acid-requiring mutant from Escherichia coli. J Bacteriol. 1967 Apr;93(4):1473–1474. doi: 10.1128/jb.93.4.1473-1474.1967. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Yanisch-Perron C., Vieira J., Messing J. Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene. 1985;33(1):103–119. doi: 10.1016/0378-1119(85)90120-9. [DOI] [PubMed] [Google Scholar]
  49. Zalkin H., Ebbole D. J. Organization and regulation of genes encoding biosynthetic enzymes in Bacillus subtilis. J Biol Chem. 1988 Feb 5;263(4):1595–1598. [PubMed] [Google Scholar]
  50. Zeigler D. R., Dean D. H. Orientation of genes in the Bacillus subtilis chromosome. Genetics. 1990 Aug;125(4):703–708. doi: 10.1093/genetics/125.4.703. [DOI] [PMC free article] [PubMed] [Google Scholar]

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