Productive folding of metallo-b-lactamases in the periplasm of Gram-negative bacteria

BRAMBILLA L; MORAN BARRIO J; VIALE AM.

Carlos Paz

Congreso; XLIV Reunión Anual SAIB; 2008

Sociedad Argentina de Investigación en Bioquímica y Biología Molecular

Periplasmic proteins are synthesized in the cytoplasm and acquire thier native structure in the final destination. While cytoplasmic chaperones have been well characterized, much less is known about periplasmic folding assistants. We studied here by genetic methods the role of proposed periplasmic chaperones in the biogenesis of the metallo-b-lactamase (MbL) GOB. We observed that the production of the GOB precursor in Escherichia coli confers b-lactam resistance, but that deletion mutants in putative periplasmic chaperones such as DsbA, DsbC, DsbG, PpiA, PpiD, SurA, FkpA or DegP did not affect resistance levels indicating that these proteins do not participate individually in GOB biogenesis. Salmonella enterica insertion mutantas were generated by MudJ transposition, and clones with diminished b-lactam antibiotic resistance were isolated. Sequence analysis of a particular mutant called RDC, showed disruption of phsB, a gene part of the phsABC operon that encodes periplasmic thiosulfate reductase responsible for H2S production during anaerobic respiration. Bioquemical assays indicated that this mutant still generate H2S, suggesting that PhsB is not essential for this function. Based on these observations, we propose that this redox protein is implicated in the productive folding of metalloenzymes such as MbLs in the bacterial periplasm.