Reversion rates in a leuB auxotroph of Escherichia coli K-12 correlate with ppGpp levels during exponential growth

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Abstract
Two isogenic strains of Escherichia coli K-12 differing only in relA, as well as two spoT transductants of the relA- strain, were examined with respect to ppGpp levels and reversion rates of a leuB- allele under nine different conditions. A positive correlation was established between reversion rates and the steady-state concentration of ppGpp during exponential growth. The leuB genes from two leuB- strains (isogenic except for relA) were cloned and sequenced and found to contain a single mutation, namely, a C-to-T transition at nucleotide 857. This mutation resulted in a serine-to-leucine substitution at amino acid residue 286 of the LeuB protein. PCR products that encompassed the leuB lesion were generated from 53 revertants and then sequenced. Of these revertants, 36 were found to contain nucleotide substitutions that would result in a serine (wild type), valine or methionine at amino acid residue 286 of LeuB, and nearly all of them exhibited generation times similar to wild type. Seventeen of the analysed revertants were found to be suppressors that retained the encoded leucine at residue 286. The majority of the suppressor mutants exhibited generation times that were significantly longer than wild type.