Abstract
Previous studies have shown that narL+ is required for nitrate regulation of anaerobic respiratory enzyme synthesis, including formate dehydrogenase-N, nitrate reductase, and fumarate reductase. Insertions in the closely linked narX gene decrease, but do not abolish, nitrate regulation of anaerobic enzyme synthesis. Analysis of sequence similarities suggests that NarX and NarL comprise a two-component regulatory pair. We constructed lacZ operon and gene fusions to investigate the operon structure of narXL. We found evidence for a complex operon with at least two promoters; PXL-narX-PL-narL. We also investigated the role of NarX in nitrate regulation of anaerobic respiratory enzyme synthesis by constructing nonpolar loss of function narX alleles. These deletions were studied on narL+ lambda specialized transducing bacteriophage. The narX deletions had no effect on nitrate regulation in delta (narXL) strains. This finding suggest that the subtle effects of previously studied narX insertions are due to decreased expression of narL and that narX+ is not essential for normal nitrate regulation. The role of NarX in nitrate regulation remains to be determined.
Full text
PDF









Images in this article
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Arps P. J., Winkler M. E. Structural analysis of the Escherichia coli K-12 hisT operon by using a kanamycin resistance cassette. J Bacteriol. 1987 Mar;169(3):1061–1070. doi: 10.1128/jb.169.3.1061-1070.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Chamberlain J. P. Fluorographic detection of radioactivity in polyacrylamide gels with the water-soluble fluor, sodium salicylate. Anal Biochem. 1979 Sep 15;98(1):132–135. doi: 10.1016/0003-2697(79)90716-4. [DOI] [PubMed] [Google Scholar]
- Chippaux M., Bonnefoy-Orth V., Ratouchniak J., Pascal M. C. Operon fusions in the nitrate reductase operon and study of the control gene nir R in Escherichia coli. Mol Gen Genet. 1981;182(3):477–479. doi: 10.1007/BF00293938. [DOI] [PubMed] [Google Scholar]
- Ciampi M. S., Roth J. R. Polarity effects in the hisG gene of salmonella require a site within the coding sequence. Genetics. 1988 Feb;118(2):193–202. doi: 10.1093/genetics/118.2.193. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Cotter P. A., Gunsalus R. P. Oxygen, nitrate, and molybdenum regulation of dmsABC gene expression in Escherichia coli. J Bacteriol. 1989 Jul;171(7):3817–3823. doi: 10.1128/jb.171.7.3817-3823.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Enoch H. G., Lester R. L. The role of a novel cytochrome b-containing nitrate reductase and quinone in the in vitro reconstruction of formate-nitrate reductase activity of E. coli. Biochem Biophys Res Commun. 1974 Dec 23;61(4):1234–1241. doi: 10.1016/s0006-291x(74)80416-x. [DOI] [PubMed] [Google Scholar]
- Fellay R., Frey J., Krisch H. Interposon mutagenesis of soil and water bacteria: a family of DNA fragments designed for in vitro insertional mutagenesis of gram-negative bacteria. Gene. 1987;52(2-3):147–154. doi: 10.1016/0378-1119(87)90041-2. [DOI] [PubMed] [Google Scholar]
- Gunsalus R. P., Kalman L. V., Stewart R. R. Nucleotide sequence of the narL gene that is involved in global regulation of nitrate controlled respiratory genes of Escherichia coli. Nucleic Acids Res. 1989 Mar 11;17(5):1965–1975. doi: 10.1093/nar/17.5.1965. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Iuchi S., Lin E. C. The narL gene product activates the nitrate reductase operon and represses the fumarate reductase and trimethylamine N-oxide reductase operons in Escherichia coli. Proc Natl Acad Sci U S A. 1987 Jun;84(11):3901–3905. doi: 10.1073/pnas.84.11.3901. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Jones H. M., Gunsalus R. P. Transcription of the Escherichia coli fumarate reductase genes (frdABCD) and their coordinate regulation by oxygen, nitrate, and fumarate. J Bacteriol. 1985 Dec;164(3):1100–1109. doi: 10.1128/jb.164.3.1100-1109.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kalman L. V., Gunsalus R. P. Identification of a second gene involved in global regulation of fumarate reductase and other nitrate-controlled genes for anaerobic respiration in Escherichia coli. J Bacteriol. 1989 Jul;171(7):3810–3816. doi: 10.1128/jb.171.7.3810-3816.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Lambden P. R., Guest J. R. Mutants of Escherichia coli K12 unable to use fumarate as an anaerobic electron acceptor. J Gen Microbiol. 1976 Dec;97(2):145–160. doi: 10.1099/00221287-97-2-145. [DOI] [PubMed] [Google Scholar]
- Li S. F., DeMoss J. A. Location of sequences in the nar promoter of Escherichia coli required for regulation by Fnr and NarL. J Biol Chem. 1988 Sep 25;263(27):13700–13705. [PubMed] [Google Scholar]
- Li S. F., DeMoss J. A. Promoter region of the nar operon of Escherichia coli: nucleotide sequence and transcription initiation signals. J Bacteriol. 1987 Oct;169(10):4614–4620. doi: 10.1128/jb.169.10.4614-4620.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Nohno T., Noji S., Taniguchi S., Saito T. The narX and narL genes encoding the nitrate-sensing regulators of Escherichia coli are homologous to a family of prokaryotic two-component regulatory genes. Nucleic Acids Res. 1989 Apr 25;17(8):2947–2957. doi: 10.1093/nar/17.8.2947. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Prentki P., Krisch H. M. In vitro insertional mutagenesis with a selectable DNA fragment. Gene. 1984 Sep;29(3):303–313. doi: 10.1016/0378-1119(84)90059-3. [DOI] [PubMed] [Google Scholar]
- Ruiz-Herrera J., DeMoss J. A. Nitrate reductase complex of Escherichia coli K-12: participation of specific formate dehydrogenase and cytochrome b1 components in nitrate reduction. J Bacteriol. 1969 Sep;99(3):720–729. doi: 10.1128/jb.99.3.720-729.1969. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Sawers G., Böck A. Anaerobic regulation of pyruvate formate-lyase from Escherichia coli K-12. J Bacteriol. 1988 Nov;170(11):5330–5336. doi: 10.1128/jb.170.11.5330-5336.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Silhavy T. J., Beckwith J. R. Uses of lac fusions for the study of biological problems. Microbiol Rev. 1985 Dec;49(4):398–418. doi: 10.1128/mr.49.4.398-418.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Simons R. W., Houman F., Kleckner N. Improved single and multicopy lac-based cloning vectors for protein and operon fusions. Gene. 1987;53(1):85–96. doi: 10.1016/0378-1119(87)90095-3. [DOI] [PubMed] [Google Scholar]
- Stewart G. S., Lubinsky-Mink S., Jackson C. G., Cassel A., Kuhn J. pHG165: a pBR322 copy number derivative of pUC8 for cloning and expression. Plasmid. 1986 May;15(3):172–181. doi: 10.1016/0147-619x(86)90035-1. [DOI] [PubMed] [Google Scholar]
- Stewart V., Berg B. L. Influence of nar (nitrate reductase) genes on nitrate inhibition of formate-hydrogen lyase and fumarate reductase gene expression in Escherichia coli K-12. J Bacteriol. 1988 Oct;170(10):4437–4444. doi: 10.1128/jb.170.10.4437-4444.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Stewart V. Nitrate respiration in relation to facultative metabolism in enterobacteria. Microbiol Rev. 1988 Jun;52(2):190–232. doi: 10.1128/mr.52.2.190-232.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Stewart V., Parales J., Jr Identification and expression of genes narL and narX of the nar (nitrate reductase) locus in Escherichia coli K-12. J Bacteriol. 1988 Apr;170(4):1589–1597. doi: 10.1128/jb.170.4.1589-1597.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Stewart V., Parales J., Jr, Merkel S. M. Structure of genes narL and narX of the nar (nitrate reductase) locus in Escherichia coli K-12. J Bacteriol. 1989 Apr;171(4):2229–2234. doi: 10.1128/jb.171.4.2229-2234.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Stewart V. Requirement of Fnr and NarL functions for nitrate reductase expression in Escherichia coli K-12. J Bacteriol. 1982 Sep;151(3):1320–1325. doi: 10.1128/jb.151.3.1320-1325.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Stewart V., Yanofsky C. Evidence for transcription antitermination control of tryptophanase operon expression in Escherichia coli K-12. J Bacteriol. 1985 Nov;164(2):731–740. doi: 10.1128/jb.164.2.731-740.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Stewart V., Yanofsky C. Role of leader peptide synthesis in tryptophanase operon expression in Escherichia coli K-12. J Bacteriol. 1986 Jul;167(1):383–386. doi: 10.1128/jb.167.1.383-386.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Stock J. B., Ninfa A. J., Stock A. M. Protein phosphorylation and regulation of adaptive responses in bacteria. Microbiol Rev. 1989 Dec;53(4):450–490. doi: 10.1128/mr.53.4.450-490.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Tabor S., Richardson C. C. A bacteriophage T7 RNA polymerase/promoter system for controlled exclusive expression of specific genes. Proc Natl Acad Sci U S A. 1985 Feb;82(4):1074–1078. doi: 10.1073/pnas.82.4.1074. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Vandeyar M. A., Weiner M. P., Hutton C. J., Batt C. A. A simple and rapid method for the selection of oligodeoxynucleotide-directed mutants. Gene. 1988 May 15;65(1):129–133. doi: 10.1016/0378-1119(88)90425-8. [DOI] [PubMed] [Google Scholar]
- Wanner B. L., Wilmes M. R., Hunter E. Molecular cloning of the wild-type phoM operon in Escherichia coli K-12. J Bacteriol. 1988 Jan;170(1):279–288. doi: 10.1128/jb.170.1.279-288.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Winans S. C., Elledge S. J., Krueger J. H., Walker G. C. Site-directed insertion and deletion mutagenesis with cloned fragments in Escherichia coli. J Bacteriol. 1985 Mar;161(3):1219–1221. doi: 10.1128/jb.161.3.1219-1221.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]