The physical role of the proteases from a marine bacterium, Alteromonas sp. Strain 0-7

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Within the extracellularproteases from the marine bacterium, Alteromonassp.strain O-7, AprIV is involved in the chitinolytic system. AprIV and chitinases (ChiA.ChiB,ChiC, and ChiD) were induced in the presence of N-acetylglucosamine (GlcNAc). To clarify the regulation of AprIV at the gene level, N-terminal amino acid sequence of the protein binding the upstream region of aprIV gene was determined. The result suggest that the regulatory protein is an anoxic redox control protein (ArcA), which is classified into the response regulator of ArcB/ArcA two-component signal transduction system. Thus, we cloned the areA and arcB gene of the strain, and expression plasmids ArcA and ArcB lacking the N-terminal membrane binding region (ArcBAN) were constructed by using the GST fusion protein expression vector pGEX-6P-1. The ArcA protein was transphosphorylated by the GstArcBΔN protein as a sensor kinase. Gel retardation assay and BIAcore analysis demonstrated that phosphorylated ArcA (ArcA-P) more tightly bound to the upstream region of aprIV and chiD genes compared to those of chiA,chiB,chiC, and cpb1 genes. These results suggest that ArcB/ArcA two-component signal transduction system regulates the expression of genes involved in chitin degradation system of the strain.