Conditional Mutation of an Essential Putative Glycoprotease Eliminates Autolysis in Staphylococcus aureus

Department of Veterinary and Biomedical Sciences, College of Veterinary Medicine, University of Minnesota. 1971 Commonwealth Ave. St. Paul. MN 55108, USA; Department of Pathology and Microbiology, University of Nebraska Medical Center, Omaha, NE 68198-6495, USA; Laboratory of Microbial Biofilms, Instituto de Agrobiotecnologia, Universidad Publica de Navarra, Pamplona-31006, Spain

^* To whom correspondence should be addressed. Email: jixxx002{at}umn.edu.

	Abstract

Our previous studies have demonstrated that a Staphylococcus aureus putative glycoprotease (Gcp) is essential for bacterial survival, indicating that Gcp may be a novel target for developing antibacterial agents. However, the biological function of Gcp is unclear. In order to elucidate the reason why Gcp is required for growth, we examined the role of Gcp in bacterial autolysis, which is an important biological process for bacterial growth. Using both a Pspac-regulated gcp expression strain and a TetR-regulated gcp antisense expression strain, we found that the down-regulation of gcp expression can effectively inhibit Triton X-100-induced lysis, eliminate penicillin- and vancomycin-caused cell lysis, and dramatically increase tolerance to hydrolases. Moreover, we determined whether resistance to lysis is due to a defect in murein hydrolase activity using a zymogram analysis. The results showed that the cell lysate of down-regulated gcp expression mutant displayed several bands of decreased murein hydrolytic activities. Furthermore, we explored the potential mechanism of Gcp's involvement in autolysis, and demonstrated that Gcp may function independently from several key autolysins (Atl, LytM and LytN) and regulators (ArlRS, Mgr/Rat, and CidA). Taken together, the above results indicate that the essential Gcp is involved in the modification of substrates of murein hydrolases, as well as regulation of expression and/or activity of some murein hydrolases, which, in turn, may play important roles in bacterial viability.