Cloning and characterization of the gene encoding the PepF endopeptidase from the aquatic bacterium Caulobacter crescentus

Abstract

The metallopeptidases have a very important role in bacteria, being involved in several processes that rely on protein turnover, such as nutrition, degradation of signal peptides, protein localization and virulence. We have cloned and characterized the gene of the metalloendopeptidase PepF from the aquatic bacterium Caulobacter crescentus. The gene upstream of pepF (orf1) encodes a conserved hypothetical protein found in Mycobacterium and Streptomyces. pepF is co-transcribed with the gene downstream (orf3), which encodes a protein that belongs to the ABC1 protein kinase family, suggesting that these two proteins may share a common function in the cell. The C. crescentus PepF protein possesses the conserved HEXGH motif present in zinc binding domains of PepF homologs. Disruption of the pepF gene by insertion of a vector sequence did not produced any growth defect, but the mutant strain possesses only 30% of the specific activity of endopeptidases present in the wild type strain. Deletions and point mutations in the regulatory region showed that there are two putative promoter regions, and the operon expression is independent of the transcription regulator CtrA. The results indicate that PepF is not essential for either growth or development of this bacterium using peptides as the sole carbon source, suggesting that other peptidases can be sharing this function.