1887

Abstract

A  : :  reporter was constructed in strains Newman and 8325-4, whereby the level of tetracycline resistance reflected the activity of the promoter. Wild-type strains carrying a single copy of this construct exhibited a low level of tetracycline resistance, suggesting that the promoter is weak. Spontaneous mutants that grew at higher tetracycline concentrations were isolated. Some were due to point mutations in the promoter that led to increased expression of the gene. The promoter was identified by primer extension analysis and −35 and −10 elements were assigned. The promoter regions from the tetracycline-resistant mutants were sequenced and several had base changes within or adjacent to the −35 box. Three created the consensus −35 sequence TTGACA. The mutant promoters were fused to . β-Galactosidase activity was six- to ninefold higher in the mutant strains compared to the wild-type. The wild-type gene was placed under the control of the mutant promoters. ClfB expression was higher than the corresponding wild-type strains and the protein was present on bacteria from the stationary phase instead of being confined to the exponential phase. Therefore, mutations in the promoter that cause changes in the −35 region produce a stronger promoter that is capable of increased transcription and, as a result, increased expression of ClfB.

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2003-01-01
2020-02-28
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