Osmoregulation in Escherichia coli: complementation analysis and gene-protein relationships in the proU locus

Abstract

The proU locus in Escherichia coli encodes an osmotically inducible transport system for two substrates, glycine betaine and L-proline, whose intracellular accumulation represents an important component in the physiology of osmoregulation. Several osmoresponsive proU::lac mutants were isolated and tested for complementation with plasmids carrying different functional regions of proU. Three classes of mutations were identified which were physically mapped to distinct regions of DNA from this locus. Tn1000-insertion mutagenesis of cloned proU DNA also yielded three phenotypic classes of mutations whose physical distribution approximately corresponded with those of the chromosomal mutants above. Three proteins, of Mr 44,000, 35,000, and 33,000, were shown to be products of proU, and the last of these was localized to the periplasmic space. The data indicate that proU is an operon with three genes, designated in order proV, proW, and proX, encoding respectively the gene products above. All three genes were shown to be necessary for exhibition of the proU-mediated osmoprotective effects of both glycine betaine and L-proline in E. coli.