PMID:2066339
| Citation |
Egan, SM and Stewart, V (1991) Mutational analysis of nitrate regulatory gene narL in Escherichia coli K-12. J. Bacteriol. 173:4424-32 |
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| Abstract |
The narL gene product, NarL, is the nitrate-responsive regulator of anaerobic respiratory gene expression. We used genetic analysis of narL mutants to better understand the mechanism of NarL-mediated gene regulation. We selected and analyzed seven nitrate-independent narL mutants. Each of three independent, strongly constitutive mutants had changes of Val-88 to Ala. The other four mutants were weakly constitutive. The narL505(V88A) allele was largely dominant to narL+, while narX+ had a negative influence on its constitutive phenotype, suggesting that NarX may play a negative role in nitrate regulation. We also constructed two narL mutations that are analogous to previously characterized constitutive degU alleles. The first, narL503(H15L), was a recessive null allele. The second, narL504(D110K), functioned essentially as wild type but was dependent on narX+ for full activity. We changed Asp-59 of NarL, which corresponds to the site of phosphorylation of other response regulators, to Asn. This change, narL502(D59N), was a recessive null allele, which is consistent with the hypothesis that NarL requires phosphorylation for activation. Finally, we tested the requirement for molybdate on regulation in a narL505(V88A) strain. Although narL505(V88A) conferred some nitrate-independent expression of fdnGHI (encoding formate dehydrogenase-N) in limiting molybdate, it required excess molybdate for full induction both in the absence and in the presence of nitrate. This finding suggests that narL505(V88A) did not confer molybdate-independent expression of fdnGHI. |
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| Keywords |
Amino Acid Sequence; Bacterial Proteins/genetics; Base Sequence; DNA-Binding Proteins; Escherichia coli/genetics; Escherichia coli Proteins; Gene Expression Regulation, Bacterial; Genes, Bacterial; Genes, Regulator; Genotype; Molecular Sequence Data; Mutagenesis, Site-Directed; Nitrates/metabolism; Oligonucleotide Probes; Plasmids; Sequence Homology, Nucleic Acid |
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