Skip to main content
PMC home page
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1989 Nov;171(11):5915–5921. doi: 10.1128/jb.171.11.5915-5921.1989

Genetic organization and sequence of the Pseudomonas cepacia genes for the alpha and beta subunits of protocatechuate 3,4-dioxygenase.

G J Zylstra 1, R H Olsen 1, D P Ballou 1
PMCID: PMC210454  PMID: 2808303

Abstract

The locations of the genes for the alpha and beta subunits of protocatechuate 3,4-dioxygenase (EC 1.13.11.3) on a 9.5-kilobase-pair PstI fragment cloned from the Pseudomonas cepacia DBO1 chromosome were determined. This was accomplished through the construction of several subclones into the broad-host-range cloning vectors pRO2317, pRO2320, and pRO2321. The ability of each subclone to complement mutations in protocatechuate 3,4-dioxygenase (pcaA) was tested in mutant strains derived from P. cepacia, Pseudomonas aeruginosa, and Pseudomonas putida. These complementation studies also showed that the two subunits were expressed from the same promoter. The nucleotide sequence of the region encoding for protocatechuate 3,4-dioxygenase was determined. The deduced amino acid sequence matched that determined by N-terminal analysis of regions of the isolated enzyme. Although over 400 nucleotides were sequenced before the start of the genes, no homology to known promoters was found. However, a terminator stem-loop structure was found immediately after the genes. The deduced amino acid sequence showed extensive homology with the previously determined amino acid sequence of protocatechuate 3,4-dioxygenase from another Pseudomonas species.

Full text

PDF
5921

Selected References

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

  1. Adhya S., Gottesman M. Control of transcription termination. Annu Rev Biochem. 1978;47:967–996. doi: 10.1146/annurev.bi.47.070178.004535. [DOI] [PubMed] [Google Scholar]
  2. Bagdasarian M., Timmis K. N. Host: vector systems for gene cloning in Pseudomonas. Curr Top Microbiol Immunol. 1982;96:47–67. doi: 10.1007/978-3-642-68315-2_4. [DOI] [PubMed] [Google Scholar]
  3. Ballard R. W., Palleroni N. J., Doudoroff M., Stanier R. Y., Mandel M. Taxonomy of the aerobic pseudomonads: Pseudomonas cepacia, P. marginata, P. alliicola and P. caryophylli. J Gen Microbiol. 1970 Feb;60(2):199–214. doi: 10.1099/00221287-60-2-199. [DOI] [PubMed] [Google Scholar]
  4. 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]
  5. Boyer H. W., Roulland-Dussoix D. A complementation analysis of the restriction and modification of DNA in Escherichia coli. J Mol Biol. 1969 May 14;41(3):459–472. doi: 10.1016/0022-2836(69)90288-5. [DOI] [PubMed] [Google Scholar]
  6. Brown N. L., Ford S. J., Pridmore R. D., Fritzinger D. C. Nucleotide sequence of a gene from the Pseudomonas transposon Tn501 encoding mercuric reductase. Biochemistry. 1983 Aug 16;22(17):4089–4095. doi: 10.1021/bi00286a015. [DOI] [PubMed] [Google Scholar]
  7. Bull C., Ballou D. P., Otsuka S. The reaction of oxygen with protocatechuate 3,4-dioxygenase from Pseudomonas putida. Characterization of a new oxygenated intermediate. J Biol Chem. 1981 Dec 25;256(24):12681–12686. [PubMed] [Google Scholar]
  8. Bull C., Ballou D. P. Purification and properties of protocatechuate 3,4-dioxygenase from Pseudomonas putida. A new iron to subunit stoichiometry. J Biol Chem. 1981 Dec 25;256(24):12673–12680. [PubMed] [Google Scholar]
  9. Bull C., Ballou D. P., Salmeen I. Raman spectrum of protocatechuate dioxygenase from Pseudomonas putida. New low frequency bands. Biochem Biophys Res Commun. 1979 Apr 13;87(3):836–841. doi: 10.1016/0006-291x(79)92033-3. [DOI] [PubMed] [Google Scholar]
  10. Bull C., Ballou D. P., Salmeen I. Raman spectrum of protocatechuate dioxygenase from Pseudomonas putida. New low frequency bands. Biochem Biophys Res Commun. 1979 Apr 13;87(3):836–841. doi: 10.1016/0006-291x(79)92033-3. [DOI] [PubMed] [Google Scholar]
  11. Crawford I. P., Eberly L. Structure and regulation of the anthranilate synthase genes in Pseudomonas aeruginosa: I. Sequence of trpG encoding the glutamine amidotransferase subunit. Mol Biol Evol. 1986 Sep;3(5):436–448. doi: 10.1093/oxfordjournals.molbev.a040408. [DOI] [PubMed] [Google Scholar]
  12. Cuskey S. M., Peccoraro V., Olsen R. H. Initial catabolism of aromatic biogenic amines by Pseudomonas aeruginosa PAO: pathway description, mapping of mutations, and cloning of essential genes. J Bacteriol. 1987 Jun;169(6):2398–2404. doi: 10.1128/jb.169.6.2398-2404.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Durham D. R., Stirling L. A., Ornston L. N., Perry J. J. Intergeneric evolutionary homology revealed by the study of protocatechuate 3,4-dioxygenase from Azotobacter vinelandii. Biochemistry. 1980 Jan 8;19(1):149–155. doi: 10.1021/bi00542a023. [DOI] [PubMed] [Google Scholar]
  14. Fujisawa H., Hayaishi O. Protocatechuate 3,4-dioxygenase. I. Crystallization and characterization. J Biol Chem. 1968 May 25;243(10):2673–2681. [PubMed] [Google Scholar]
  15. Garoff H., Ansorge W. Improvements of DNA sequencing gels. Anal Biochem. 1981 Aug;115(2):450–457. doi: 10.1016/0003-2697(81)90031-2. [DOI] [PubMed] [Google Scholar]
  16. Gray G. L., Smith D. H., Baldridge J. S., Harkins R. N., Vasil M. L., Chen E. Y., Heyneker H. L. Cloning, nucleotide sequence, and expression in Escherichia coli of the exotoxin A structural gene of Pseudomonas aeruginosa. Proc Natl Acad Sci U S A. 1984 May;81(9):2645–2649. doi: 10.1073/pnas.81.9.2645. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hadero A., Crawford I. P. Nucleotide sequence of the genes for tryptophan synthase in Pseudomonas aeruginosa. Mol Biol Evol. 1986 May;3(3):191–204. doi: 10.1093/oxfordjournals.molbev.a040388. [DOI] [PubMed] [Google Scholar]
  18. Holloway B. W., Krishnapillai V., Morgan A. F. Chromosomal genetics of Pseudomonas. Microbiol Rev. 1979 Mar;43(1):73–102. doi: 10.1128/mr.43.1.73-102.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Hou C. T., Lillard M. O., Schwartz R. D. Protocatechuate 3, 4-dioxygenase from Acinetobacter calcoaceticus. Biochemistry. 1976 Feb 10;15(3):582–588. doi: 10.1021/bi00648a020. [DOI] [PubMed] [Google Scholar]
  20. Inouye S., Asai Y., Nakazawa A., Nakazawa T. Nucleotide sequence of a DNA segment promoting transcription in Pseudomonas putida. J Bacteriol. 1986 Jun;166(3):739–745. doi: 10.1128/jb.166.3.739-745.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Inouye S., Ebina Y., Nakazawa A., Nakazawa T. Nucleotide sequence surrounding transcription initiation site of xylABC operon on TOL plasmid of Pseudomonas putida. Proc Natl Acad Sci U S A. 1984 Mar;81(6):1688–1691. doi: 10.1073/pnas.81.6.1688. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. 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]
  23. Iwaki M., Kagamiyama H., Nozaki M. The primary structure of the beta-subunit of protocatechuate 3,4-dioxygenase from Pseudomonas aeruginosa. Arch Biochem Biophys. 1981 Aug;210(1):210–223. doi: 10.1016/0003-9861(81)90182-x. [DOI] [PubMed] [Google Scholar]
  24. Kohlmiller N. A., Howard J. B. The primary structure of the alpha subunit of protocatechuate 3,4-dioxygenase. II. Isolation and sequence of overlap peptides and complete sequence. J Biol Chem. 1979 Aug 10;254(15):7309–7315. [PubMed] [Google Scholar]
  25. Ludwig M. L., Weber L. D., Ballou D. P. Characterization of crystals of protocatechuate 3,4-dioxygenase from Pseudomonas cepacia. J Biol Chem. 1984 Dec 10;259(23):14840–14842. [PubMed] [Google Scholar]
  26. Maeda K., Kneale G. G., Tsugita A., Short N. J., Perham R. N., Hill D. F., Petersen G. B. The DNA-binding protein of Pf1 filamentous bacteriophage: amino-acid sequence and structure of the gene. EMBO J. 1982;1(2):255–261. doi: 10.1002/j.1460-2075.1982.tb01156.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Matsuda A., Komatsu K. I. Molecular cloning and structure of the gene for 7 beta-(4-carboxybutanamido)cephalosporanic acid acylase from a Pseudomonas strain. J Bacteriol. 1985 Sep;163(3):1222–1228. doi: 10.1128/jb.163.3.1222-1228.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Mercer A. A., Loutit J. S. Transformation and transfection of Pseudomonas aeruginosa: effects of metal ions. J Bacteriol. 1979 Oct;140(1):37–42. doi: 10.1128/jb.140.1.37-42.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Mermod N., Lehrbach P. R., Reineke W., Timmis K. N. Transcription of the TOL plasmid toluate catabolic pathway operon of Pseudomonas putida is determined by a pair of co-ordinately and positively regulated overlapping promoters. EMBO J. 1984 Nov;3(11):2461–2466. doi: 10.1002/j.1460-2075.1984.tb02156.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Minton N. P., Atkinson T., Bruton C. J., Sherwood R. F. The complete nucleotide sequence of the Pseudomonas gene coding for carboxypeptidase G2. Gene. 1984 Nov;31(1-3):31–38. doi: 10.1016/0378-1119(84)90192-6. [DOI] [PubMed] [Google Scholar]
  31. Misra T. K., Brown N. L., Fritzinger D. C., Pridmore R. D., Barnes W. M., Haberstroh L., Silver S. Mercuric ion-resistance operons of plasmid R100 and transposon Tn501: the beginning of the operon including the regulatory region and the first two structural genes. Proc Natl Acad Sci U S A. 1984 Oct;81(19):5975–5979. doi: 10.1073/pnas.81.19.5975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Nakai C., Kagamiyama H., Nozaki M., Nakazawa T., Inouye S., Ebina Y., Nakazawa A. Complete nucleotide sequence of the metapyrocatechase gene on the TOI plasmid of Pseudomonas putida mt-2. J Biol Chem. 1983 Mar 10;258(5):2923–2928. [PubMed] [Google Scholar]
  33. Olsen R. H., DeBusscher G., McCombie W. R. Development of broad-host-range vectors and gene banks: self-cloning of the Pseudomonas aeruginosa PAO chromosome. J Bacteriol. 1982 Apr;150(1):60–69. doi: 10.1128/jb.150.1.60-69.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Olsen R. H., Shipley P. Host range and properties of the Pseudomonas aeruginosa R factor R1822. J Bacteriol. 1973 Feb;113(2):772–780. doi: 10.1128/jb.113.2.772-780.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Platt T. Transcription termination and the regulation of gene expression. Annu Rev Biochem. 1986;55:339–372. doi: 10.1146/annurev.bi.55.070186.002011. [DOI] [PubMed] [Google Scholar]
  36. Rosenberg M., Court D. Regulatory sequences involved in the promotion and termination of RNA transcription. Annu Rev Genet. 1979;13:319–353. doi: 10.1146/annurev.ge.13.120179.001535. [DOI] [PubMed] [Google Scholar]
  37. 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]
  38. Sastry P. A., Finlay B. B., Pasloske B. L., Paranchych W., Pearlstone J. R., Smillie L. B. Comparative studies of the amino acid and nucleotide sequences of pilin derived from Pseudomonas aeruginosa PAK and PAO. J Bacteriol. 1985 Nov;164(2):571–577. doi: 10.1128/jb.164.2.571-577.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Shine J., Dalgarno L. Determinant of cistron specificity in bacterial ribosomes. Nature. 1975 Mar 6;254(5495):34–38. doi: 10.1038/254034a0. [DOI] [PubMed] [Google Scholar]
  40. Stanier R. Y., Palleroni N. J., Doudoroff M. The aerobic pseudomonads: a taxonomic study. J Gen Microbiol. 1966 May;43(2):159–271. doi: 10.1099/00221287-43-2-159. [DOI] [PubMed] [Google Scholar]
  41. Unger B. P., Gunsalus I. C., Sligar S. G. Nucleotide sequence of the Pseudomonas putida cytochrome P-450cam gene and its expression in Escherichia coli. J Biol Chem. 1986 Jan 25;261(3):1158–1163. [PubMed] [Google Scholar]
  42. VOGEL H. J., BONNER D. M. Acetylornithinase of Escherichia coli: partial purification and some properties. J Biol Chem. 1956 Jan;218(1):97–106. [PubMed] [Google Scholar]
  43. Walsh T. A., Ballou D. P. Halogenated protocatechuates as substrates for protocatechuate dioxygenase from Pseudomonas cepacia. J Biol Chem. 1983 Dec 10;258(23):14413–14421. [PubMed] [Google Scholar]
  44. Whittaker J. W., Lipscomb J. D., Kent T. A., Münck E. Brevibacterium fuscum protocatechuate 3,4-dioxygenase. Purification, crystallization, and characterization. J Biol Chem. 1984 Apr 10;259(7):4466–4475. [PubMed] [Google Scholar]
  45. Yoshida R., Hori K., Fujiwara M., Saeki Y., Kagamiyama H. Nonidentical subunits of protocatechuate 3,4-dioxygenase. Biochemistry. 1976 Sep 7;15(18):4048–4053. doi: 10.1021/bi00663a020. [DOI] [PubMed] [Google Scholar]
  46. Zylstra G. J., Olsen R. H., Ballou D. P. Cloning, expression, and regulation of the Pseudomonas cepacia protocatechuate 3,4-dioxygenase genes. J Bacteriol. 1989 Nov;171(11):5907–5914. doi: 10.1128/jb.171.11.5907-5914.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES