PubMed · 6247780
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T Bergan. 1980-01-30. [Mecillinam].. https://pubmed.ncbi.nlm.nih.gov/6247780/
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Escherichia coli strains partially induced for the stringent response are resistant to mecillinam, a beta-lactam antibiotic which specifically inactivates penicillin-binding protein 2, the key enzyme determining cell shape. We present evidence that mecillinam resistance occurs whenever the intracellular concentration of the nucleotide ppGpp (guanosine 3'-diphosphate 5'-diphosphate), the effector of the stringent response, exceeds a threshold level. First, the ppGpp concentration was higher in a mecillinam-resistant mutant than in closely related sensitive strains. Second, the ppGpp pool was controlled by means of a plasmid carrying a ptac-relA' gene coding for a hyperactive (p)ppGpp synthetase, RelA'; increasing the ppGpp pool by varying the concentration of lac operon inducer IPTG resulted in a sharp threshold ppGpp concentration, above which cells were mecillinam resistant. Third, the ppGpp pool was increased by using poor media; again, at the lowest growth rate studied, the cells were mecillinam resistant. In all experiments, cells with a ppGpp concentration above 140 pmoles/A600 were mecillinam resistant whereas those with lower concentrations were sensitive. We discuss a possible role for ppGpp as transcriptional activator of cell division genes whose products seem to become limiting in the presence of mecillinam, when cells form large spheres. We confirmed the well-known inverse correlation between growth rate and ppGpp concentration but, surprisingly, for a given growth rate, the ppGpp concentration was lower in poor medium than in richer medium in which RelA' is induced. We conclude that, for E. coli growing in poor media, the concentration of the nucleotide ppGpp is not the major growth rate determinant.
The Escherichia coli strain known as GC2553, FB8, UTH1038, or K12S (Luria), considered an F- lambda- wild-type strain, is shown here to carry a cryptic mutation, ftsR1, causing nonlethal filamentation during exponential growth in Luria-Bertani (LB) broth at 42 degrees C and the inability to grow in salt-free LB broth at 42 degrees C. The ftsR1 mutation is completely suppressed in genetic backgrounds which increase RelA-dependent synthesis of the nucleotide ppGpp, i.e., argS201 (Mecr) and alaS21 (Mecr) mutations, affecting aminoacyl-tRNA synthetases, or the presence of a plac-relA' plasmid. These backgrounds also confer resistance in LB broth to the beta-lactam mecillinam, an antibiotic which specifically inhibits penicillin-binding protein 2 and, in wild-type cells, causes an indirect block in cell division. Furthermore, the ftsR1 mutant (but not an isogenic ftsR+ strain) is sensitive to mecillinam in minimal glucose medium at 37 degrees C. Since the division block caused by mecillinam can be overcome by overproduction of the cell division protein FtsZ, we tested the effect of plasmid pZAQ (carrying the ftsZ, ftsA, and ftsQ genes) on the ftsR1 mutant; it suppressed the filamentation in LB broth and the mecillinam sensitivity on minimal glucose medium at 37 degrees C but not the growth defect in salt-free LB broth at 42 degrees C. Genetic analysis indicated that the full phenotype of the ftsR1 mutant is due to a single mutation in the rpoB gene (90 min), coding for the beta subunit of RNA polymerase; we call this allele rpoB369(Fts). We propose that the rpoB369(Fts) mutation alters the specificity of the polymerase and that the mutant enzyme can recover normal activity in the presence of high salt concentrations or via interaction with the nucleotide ppGpp.
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