Skip to main content
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1997 Apr;179(8):2540–2550. doi: 10.1128/jb.179.8.2540-2550.1997

Xanthine metabolism in Bacillus subtilis: characterization of the xpt-pbuX operon and evidence for purine- and nitrogen-controlled expression of genes involved in xanthine salvage and catabolism.

L C Christiansen 1, S Schou 1, P Nygaard 1, H H Saxild 1
PMCID: PMC179002  PMID: 9098051

Abstract

The xpt and pbuX genes from Bacillus subtilis were cloned, and their nucleotide sequences were determined. The xpt gene encodes a specific xanthine phosphoribosyltransferase, and the pbuX gene encodes a xanthine-specific purine permease. The genes have overlapping coding regions, and Northern (RNA) blot analysis indicated an operon organization. The translation of the second gene, pbuX, was strongly dependent on the translation of the first gene, xpt. Expression of the operon was repressed by purines, and the effector molecules appear to be hypoxanthine and guanine. When hypoxanthine and guanine were added together, a 160-fold repression was observed. The regulation of expression was at the level of transcription, and we propose that a transcription termination-antitermination control mechanism similar to the one suggested for the regulation of the purine biosynthesis operon exists. The expression of the xpt-pbuX operon was reduced when hypoxanthine served as the sole nitrogen source. Under these conditions, the level of the hypoxanthine- and xanthine-degrading enzyme, xanthine dehydrogenase, was induced more than 80-fold. The xanthine dehydrogenase level was completely derepressed in a glnA (glutamine synthetase) genetic background. Although the regulation of the expression of the xpt-pbuX operon was found to be affected by the nitrogen source, it was normal in a glnA mutant strain. This result suggests the existence of different signalling pathways for repression of the transcription of the xpt-pbuX operon and the induction of xanthine dehydrogenase.

Full Text

The Full Text of this article is available as a PDF (1,005.7 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Altschul S. F., Gish W., Miller W., Myers E. W., Lipman D. J. Basic local alignment search tool. J Mol Biol. 1990 Oct 5;215(3):403–410. doi: 10.1016/S0022-2836(05)80360-2. [DOI] [PubMed] [Google Scholar]
  2. Andersen P. S., Frees D., Fast R., Mygind B. Uracil uptake in Escherichia coli K-12: isolation of uraA mutants and cloning of the gene. J Bacteriol. 1995 Apr;177(8):2008–2013. doi: 10.1128/jb.177.8.2008-2013.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Atkinson M. R., Fisher S. H. Identification of genes and gene products whose expression is activated during nitrogen-limited growth in Bacillus subtilis. J Bacteriol. 1991 Jan;173(1):23–27. doi: 10.1128/jb.173.1.23-27.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Beaman T. C., Hitchins A. D., Ochi K., Vasantha N., Endo T., Freese E. Specificity and control of uptake of purines and other compounds in Bacillus subtilis. J Bacteriol. 1983 Dec;156(3):1107–1117. doi: 10.1128/jb.156.3.1107-1117.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Birnboim H. C., Doly J. A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res. 1979 Nov 24;7(6):1513–1523. doi: 10.1093/nar/7.6.1513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Brown K. L., Hughes K. T. The role of anti-sigma factors in gene regulation. Mol Microbiol. 1995 May;16(3):397–404. doi: 10.1111/j.1365-2958.1995.tb02405.x. [DOI] [PubMed] [Google Scholar]
  7. Burton K. Transport of nucleic acid bases into Escherichia coli. J Gen Microbiol. 1983 Nov;129(11):3505–3513. doi: 10.1099/00221287-129-11-3505. [DOI] [PubMed] [Google Scholar]
  8. Caramori T., Calogero S., Albertini A. M., Galizzi A. Functional analysis of the outB gene of Bacillus subtilis. J Gen Microbiol. 1993 Jan;139(1):31–37. doi: 10.1099/00221287-139-1-31. [DOI] [PubMed] [Google Scholar]
  9. Danielsen S., Kilstrup M., Barilla K., Jochimsen B., Neuhard J. Characterization of the Escherichia coli codBA operon encoding cytosine permease and cytosine deaminase. Mol Microbiol. 1992 May;6(10):1335–1344. doi: 10.1111/j.1365-2958.1992.tb00854.x. [DOI] [PubMed] [Google Scholar]
  10. Diallinas G., Gorfinkiel L., Arst H. N., Jr, Cecchetto G., Scazzocchio C. Genetic and molecular characterization of a gene encoding a wide specificity purine permease of Aspergillus nidulans reveals a novel family of transporters conserved in prokaryotes and eukaryotes. J Biol Chem. 1995 Apr 14;270(15):8610–8622. doi: 10.1074/jbc.270.15.8610. [DOI] [PubMed] [Google Scholar]
  11. Ebbole D. J., Zalkin H. Cloning and characterization of a 12-gene cluster from Bacillus subtilis encoding nine enzymes for de novo purine nucleotide synthesis. J Biol Chem. 1987 Jun 15;262(17):8274–8287. [PubMed] [Google Scholar]
  12. Endo T., Uratani B., Freese E. Purine salvage pathways of Bacillus subtilis and effect of guanine on growth of GMP reductase mutants. J Bacteriol. 1983 Jul;155(1):169–179. doi: 10.1128/jb.155.1.169-179.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Errington J. A general method for fusion of the Escherichia coli lacZ gene to chromosomal genes in Bacillus subtilis. J Gen Microbiol. 1986 Nov;132(11):2953–2966. doi: 10.1099/00221287-132-11-2953. [DOI] [PubMed] [Google Scholar]
  14. Glaser P., Danchin A., Kunst F., Zuber P., Nakano M. M. Identification and isolation of a gene required for nitrate assimilation and anaerobic growth of Bacillus subtilis. J Bacteriol. 1995 Feb;177(4):1112–1115. doi: 10.1128/jb.177.4.1112-1115.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Hanahan D. Studies on transformation of Escherichia coli with plasmids. J Mol Biol. 1983 Jun 5;166(4):557–580. doi: 10.1016/s0022-2836(83)80284-8. [DOI] [PubMed] [Google Scholar]
  16. Houlberg U., Jensen K. F. Role of hypoxanthine and guanine in regulation of Salmonella typhimurium pur gene expression. J Bacteriol. 1983 Feb;153(2):837–845. doi: 10.1128/jb.153.2.837-845.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Kelley W. N., Rosenbloom F. M., Henderson J. F., Seegmiller J. E. Xanthine phosphoribosyltransferase in man: relationship to hypoxanthine-guanine phosphoribosyltransferase. Biochem Biophys Res Commun. 1967 Aug 7;28(3):340–345. doi: 10.1016/0006-291x(67)90315-4. [DOI] [PubMed] [Google Scholar]
  18. Krenitsky T. A., Neil S. M., Miller R. L. Guanine and xanthine phosphoribosyltransfer activities of Lactobacillus casei and Escherichia coli. Their relationship to hypoxanthine and adenine phosphoribosyltransfer activities. J Biol Chem. 1970 May 25;245(10):2605–2611. [PubMed] [Google Scholar]
  19. MONOD J., COHEN-BAZIRE G., COHN M. Sur la biosynthèse de la beta-galactosidase (lactase) chez Escherichia coli; la spécificité de l'induction. Biochim Biophys Acta. 1951 Nov;7(4):585–599. doi: 10.1016/0006-3002(51)90072-8. [DOI] [PubMed] [Google Scholar]
  20. Miller R. L., Adamczyk D. L., Fyfe J. A., Elion G. B. Xanthine phosphoribosyltransferase from Streptococcus faecalis. Properties and specificity. Arch Biochem Biophys. 1974 Nov;165(1):349–358. doi: 10.1016/0003-9861(74)90173-8. [DOI] [PubMed] [Google Scholar]
  21. Mulligan R. C., Berg P. Selection for animal cells that express the Escherichia coli gene coding for xanthine-guanine phosphoribosyltransferase. Proc Natl Acad Sci U S A. 1981 Apr;78(4):2072–2076. doi: 10.1073/pnas.78.4.2072. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Nilsson B., Uhlén M., Josephson S., Gatenbeck S., Philipson L. An improved positive selection plasmid vector constructed by oligonucleotide mediated mutagenesis. Nucleic Acids Res. 1983 Nov 25;11(22):8019–8030. doi: 10.1093/nar/11.22.8019. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Nygaard P., Duckert P., Saxild H. H. Role of adenine deaminase in purine salvage and nitrogen metabolism and characterization of the ade gene in Bacillus subtilis. J Bacteriol. 1996 Feb;178(3):846–853. doi: 10.1128/jb.178.3.846-853.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Rouf M. A., Lomprey R. F., Jr Degradation of uric acid by certain aerobic bacteria. J Bacteriol. 1968 Sep;96(3):617–622. doi: 10.1128/jb.96.3.617-622.1968. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. 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]
  26. Saxild H. H., Andersen L. N., Hammer K. Dra-nupC-pdp operon of Bacillus subtilis: nucleotide sequence, induction by deoxyribonucleosides, and transcriptional regulation by the deoR-encoded DeoR repressor protein. J Bacteriol. 1996 Jan;178(2):424–434. doi: 10.1128/jb.178.2.424-434.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Saxild H. H., Jacobsen J. H., Nygaard P. Functional analysis of the Bacillus subtilis purT gene encoding formate-dependent glycinamide ribonucleotide transformylase. Microbiology. 1995 Sep;141(Pt 9):2211–2218. doi: 10.1099/13500872-141-9-2211. [DOI] [PubMed] [Google Scholar]
  28. Saxild H. H., Nygaard P. Genetic and physiological characterization of Bacillus subtilis mutants resistant to purine analogs. J Bacteriol. 1987 Jul;169(7):2977–2983. doi: 10.1128/jb.169.7.2977-2983.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Shimotsu H., Kuroda M. I., Yanofsky C., Henner D. J. Novel form of transcription attenuation regulates expression the Bacillus subtilis tryptophan operon. J Bacteriol. 1986 May;166(2):461–471. doi: 10.1128/jb.166.2.461-471.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Tinoco I., Jr, Borer P. N., Dengler B., Levin M. D., Uhlenbeck O. C., Crothers D. M., Bralla J. Improved estimation of secondary structure in ribonucleic acids. Nat New Biol. 1973 Nov 14;246(150):40–41. doi: 10.1038/newbio246040a0. [DOI] [PubMed] [Google Scholar]
  31. Tuerk C., Gauss P., Thermes C., Groebe D. R., Gayle M., Guild N., Stormo G., d'Aubenton-Carafa Y., Uhlenbeck O. C., Tinoco I., Jr CUUCGG hairpins: extraordinarily stable RNA secondary structures associated with various biochemical processes. Proc Natl Acad Sci U S A. 1988 Mar;85(5):1364–1368. doi: 10.1073/pnas.85.5.1364. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Turner R. J., Lu Y., Switzer R. L. Regulation of the Bacillus subtilis pyrimidine biosynthetic (pyr) gene cluster by an autogenous transcriptional attenuation mechanism. J Bacteriol. 1994 Jun;176(12):3708–3722. doi: 10.1128/jb.176.12.3708-3722.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Vogels G. D., Van der Drift C. Degradation of purines and pyrimidines by microorganisms. Bacteriol Rev. 1976 Jun;40(2):403–468. doi: 10.1128/br.40.2.403-468.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Weng M., Nagy P. L., Zalkin H. Identification of the Bacillus subtilis pur operon repressor. Proc Natl Acad Sci U S A. 1995 Aug 1;92(16):7455–7459. doi: 10.1073/pnas.92.16.7455. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. 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]
  36. von Heijne G. Membrane proteins: from sequence to structure. Annu Rev Biophys Biomol Struct. 1994;23:167–192. doi: 10.1146/annurev.bb.23.060194.001123. [DOI] [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES