Molecular cloning, sequence and characterization of cjsT, a putative protease from Rickettsia rickettsii

The cloning and sequencing of a gene from Rickettsia rickettsii which confers haemolytic activity on Escherichia coli strain TB1 is described. The open reading frame of the haemolysis-promoting gene, cjsT, is 1041 bp and encodes a putative protein with a molecular mass of 33 825 Da. CjsT has high sequence similarity to several bacterial proteases, particularly type IV signal peptidases. Cell lysates from an E. coli clone containing cjsT in pUC19 (pJON1) exhibited greater protease activity in functional assays than found in E. coli containing pUC19 alone. Disruption of the cjsT gene by insertional inactivation with a kanamycin cassette reduced both the protease and haemolytic activities conferred by cjsT. The protease inhibitors antipain and diisopropylfluorophosphate (DFP) both reduced the proteolytic activity of pJON1. The mechanism by which the R. rickettsii cjsT promotes haemolysis in E. coli remains unclear.