GENES INDUCED BY PH PERTURBATION ON ESCHERICHIA COLI

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Genes responsive to perturbation of external pH (an "external pH stimulon") and to perturbation of internal pH (an "internal pH stimulon") will be defined in Escherichia coli. Two distinct classes of pH-sensitive lacZ fusion strains have already been isolated: (A) strains inducible by lowering of external pH, but not by addition of a membrane-permeable weak acid at steady pH, and (B) strains inducible by weak-acid-mediate lowering of internal pH, but not by external; acidification. The fusion strains were isolated through pH screening methods devised by Slonczewski, using a Mud-fusion pool generated with MudI1734 (lac, Km), a vector constructed by M. Casadaban. Their degree of induction ranges from 10-fold to 800-fold. Previously, there are no published reports of gene fusion strains inducible by acidity, and no reports of genes induced by internal but not external perturbation of pH (Booth, 1985.) The pH-sensitive loci will be characterized with respect to genetics and biochemistry. Additional mutants will be sought which exhibit induction by alkaline perturbation of external and of internal pH. It may be that separate "stimulons" can be defined for genes responsive to alkalinization, external and internal, as well as to acidification, external and internal. The anaerobiosis response of pH-responsive gene fusion strains will be tested. Since the transmembrane pH difference (delta pH) contributes to the proton potential, one might expect some acid- inducible genes to be included in the anaerobiosis stimulon. Known anaerobiosis genes will also be tested for pH-inducibility. Two-dimensional gel electrophoresis will be perforemd to analyze proteins induced by growth in acid medium (external acid stress) or in the presence of a membrane-permeable weak acid (internal acid stress). Similar experiments will be performed to detect genes induced by alkaline stress.