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. 1992 Aug;174(16):5309–5316. doi: 10.1128/jb.174.16.5309-5316.1992

Cloning and sequencing of Escherichia coli ubiC and purification of chorismate lyase.

B P Nichols 1, J M Green 1
PMCID: PMC206367  PMID: 1644758

Abstract

In Escherichia coli, chorismate lyase catalyzes the first step in ubiquinone biosynthesis, the conversion of chorismate to 4-hydroxybenzoate. 4-Hydroxybenzoate is converted to 3-octaprenyl-4-hydroxybenzoate by 4-hydroxybenzoate octaprenyltransferase. These two enzymes are encoded by ubiC and ubiA, respectively, and have been reported to map near one another at 92 min on the E. coli chromosome. We have cloned the ubiCA gene cluster and determined the nucleotide sequence of ubiC and a portion of ubiA. The nucleotide sequence abuts with a previously determined sequence that encodes a large portion of ubiA. ubiC was localized by subcloning, and overproducing plasmids were constructed. Overexpression of ubiC allowed the purification of chorismate lyase to homogeneity, and N-terminal sequence analysis of chorismate lyase unambiguously defined the beginning of the ubiC coding region. Although chorismate lyase showed no significant amino acid sequence similarity to 4-amino-4-deoxychorismate lyase (4-amino-4-deoxychroismate----4-aminobenzoate), the product of E. coli pabC, chorismate lyase overproduction could complement the growth requirement for 4-aminobenzoate of a pabC mutant strain. Of the several enzymes that convert chorismate to intermediates of E. coli biosynthetic pathways, chorismate lyase is the last to be isolated and characterized.

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Selected References

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  1. Baker T. I., Crawford I. P. Anthranilate synthetase. Partial purification and some kinetic studies on the enzyme from Escherichia coli. J Biol Chem. 1966 Dec 10;241(23):5577–5584. [PubMed] [Google Scholar]
  2. Chepuri V., Lemieux L., Au D. C., Gennis R. B. The sequence of the cyo operon indicates substantial structural similarities between the cytochrome o ubiquinol oxidase of Escherichia coli and the aa3-type family of cytochrome c oxidases. J Biol Chem. 1990 Jul 5;265(19):11185–11192. [PubMed] [Google Scholar]
  3. Clarke L., Carbon J. A colony bank containing synthetic Col El hybrid plasmids representative of the entire E. coli genome. Cell. 1976 Sep;9(1):91–99. doi: 10.1016/0092-8674(76)90055-6. [DOI] [PubMed] [Google Scholar]
  4. Dopheide T. A., Crewther P., Davidson B. E. Chorismate mutase-prephenate dehydratase from Escherichia coli K-12. II. Kinetic properties. J Biol Chem. 1972 Jul 25;247(14):4447–4452. [PubMed] [Google Scholar]
  5. Gibson M. I., Gibson F. Preliminary studies on the isolation and metabolism of an intermediate in aromatic biosynthesis: chorismic acid. Biochem J. 1964 Feb;90(2):248–256. doi: 10.1042/bj0900248. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Goncharoff P., Nichols B. P. Nucleotide sequence of Escherichia coli pabB indicates a common evolutionary origin of p-aminobenzoate synthetase and anthranilate synthetase. J Bacteriol. 1984 Jul;159(1):57–62. doi: 10.1128/jb.159.1.57-62.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Green J. M., Merkel W. K., Nichols B. P. Characterization and sequence of Escherichia coli pabC, the gene encoding aminodeoxychorismate lyase, a pyridoxal phosphate-containing enzyme. J Bacteriol. 1992 Aug;174(16):5317–5323. doi: 10.1128/jb.174.16.5317-5323.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Green J. M., Nichols B. P. p-Aminobenzoate biosynthesis in Escherichia coli. Purification of aminodeoxychorismate lyase and cloning of pabC. J Biol Chem. 1991 Jul 15;266(20):12971–12975. [PubMed] [Google Scholar]
  9. Henikoff S. Unidirectional digestion with exonuclease III in DNA sequence analysis. Methods Enzymol. 1987;155:156–165. doi: 10.1016/0076-6879(87)55014-5. [DOI] [PubMed] [Google Scholar]
  10. Hudson G. S., Davidson B. E. Nucleotide sequence and transcription of the phenylalanine and tyrosine operons of Escherichia coli K12. J Mol Biol. 1984 Dec 25;180(4):1023–1051. doi: 10.1016/0022-2836(84)90269-9. [DOI] [PubMed] [Google Scholar]
  11. Hudson G. S., Wong V., Davidson B. E. Chorismate mutase/prephenate dehydrogenase from Escherichia coli K12: purification, characterization, and identification of a reactive cysteine. Biochemistry. 1984 Dec 4;23(25):6240–6249. doi: 10.1021/bi00320a054. [DOI] [PubMed] [Google Scholar]
  12. Kaplan J. B., Nichols B. P. Nucleotide sequence of Escherichia coli pabA and its evolutionary relationship to trp(G)D. J Mol Biol. 1983 Aug 15;168(3):451–468. doi: 10.1016/s0022-2836(83)80295-2. [DOI] [PubMed] [Google Scholar]
  13. Kohara Y., Akiyama K., Isono K. The physical map of the whole E. coli chromosome: application of a new strategy for rapid analysis and sorting of a large genomic library. Cell. 1987 Jul 31;50(3):495–508. doi: 10.1016/0092-8674(87)90503-4. [DOI] [PubMed] [Google Scholar]
  14. Kuramitsu S., Ogawa T., Ogawa H., Kagamiyama H. Branched-chain amino acid aminotransferase of Escherichia coli: nucleotide sequence of the ilvE gene and the deduced amino acid sequence. J Biochem. 1985 Apr;97(4):993–999. doi: 10.1093/oxfordjournals.jbchem.a135176. [DOI] [PubMed] [Google Scholar]
  15. Lawrence J., Cox G. B., Gibson F. Biosynthesis of ubiquinone in Escherichia coli K-12: biochemical and genetic characterization of a mutant unable to convert chorismate into 4-hydroxybenzoate. J Bacteriol. 1974 Apr;118(1):41–45. doi: 10.1128/jb.118.1.41-45.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Lightner V. A., Bell R. M., Modrich P. The DNA sequences encoding plsB and dgk loci of Escherichia coli. J Biol Chem. 1983 Sep 25;258(18):10856–10861. [PubMed] [Google Scholar]
  17. Little J. W. Isolation of recombinant plasmids and phage carrying the lexA gene of Escherichia coli K-12. Gene. 1980 Aug;10(3):237–247. doi: 10.1016/0378-1119(80)90053-0. [DOI] [PubMed] [Google Scholar]
  18. Liu J., Quinn N., Berchtold G. A., Walsh C. T. Overexpression, purification, and characterization of isochorismate synthase (EntC), the first enzyme involved in the biosynthesis of enterobactin from chorismate. Biochemistry. 1990 Feb 13;29(6):1417–1425. doi: 10.1021/bi00458a012. [DOI] [PubMed] [Google Scholar]
  19. Nichols B. P., Miozzari G. F., van Cleemput M., Bennett G. N., Yanofsky C. Nucleotide sequences of the trpG regions of Escherichia coli, Shigella dysenteriae, Salmonella typhimurium and Serratia marcescens. J Mol Biol. 1980 Oct 5;142(4):503–517. doi: 10.1016/0022-2836(80)90260-0. [DOI] [PubMed] [Google Scholar]
  20. Nichols B. P., Seibold A. M., Doktor S. Z. para-aminobenzoate synthesis from chorismate occurs in two steps. J Biol Chem. 1989 May 25;264(15):8597–8601. [PubMed] [Google Scholar]
  21. Nichols B. P., van Cleemput M., Yanofsky C. Nucleotide sequence of Escherichia coli trpE. Anthranilate synthetase component I contains no tryptophan residues. J Mol Biol. 1981 Feb 15;146(1):45–54. doi: 10.1016/0022-2836(81)90365-x. [DOI] [PubMed] [Google Scholar]
  22. Ozenberger B. A., Brickman T. J., McIntosh M. A. Nucleotide sequence of Escherichia coli isochorismate synthetase gene entC and evolutionary relationship of isochorismate synthetase and other chorismate-utilizing enzymes. J Bacteriol. 1989 Feb;171(2):775–783. doi: 10.1128/jb.171.2.775-783.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Pearson W. R., Lipman D. J. Improved tools for biological sequence comparison. Proc Natl Acad Sci U S A. 1988 Apr;85(8):2444–2448. doi: 10.1073/pnas.85.8.2444. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Ye Q. Z., Liu J., Walsh C. T. p-Aminobenzoate synthesis in Escherichia coli: purification and characterization of PabB as aminodeoxychorismate synthase and enzyme X as aminodeoxychorismate lyase. Proc Natl Acad Sci U S A. 1990 Dec;87(23):9391–9395. doi: 10.1073/pnas.87.23.9391. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Young I. G., Langman L., Luke R. K., Gibson F. Biosynthesis of the iron-transport compound enterochelin: mutants of Escherichia coli unable to synthesize 2,3-dihydroxybenzoate. J Bacteriol. 1971 Apr;106(1):51–57. doi: 10.1128/jb.106.1.51-57.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Young I. G., Leppik R. A., Hamilton J. A., Gibson F. Biochemical and genetic studies on ubiquinone biosynthesis in Escherichia coli K-12:4-hydroxybenzoate octaprenyltransferase. J Bacteriol. 1972 Apr;110(1):18–25. doi: 10.1128/jb.110.1.18-25.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]

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