• Cloning and purification of SAS0397 from community associated Staphylococcus aureus

      Wannemacher, Amber; Dannelly, Kathleen H (2012-05-21)
      Methicillin resistant Staphylococcus aureus (MRSA) is a unique bacterium that can cause painful abscesses that require surgical treatment and can be confined to the skin surface or burrow into the skin causing life-threatening infections. There are two types of MRSA: hospital associated-MRSA (HA-MRSA) and community associated-MRSA (CA-MRSA). Community associated-MRSA is acquired through direct contact with an infected individual or sharing of personal items and is often seen in locations such as athletic facilities, dormitories, and daycare centers. This study involves cloning and purification of SAS0397, a putative exported protein predicted, according to bioinformatics, to be unique to community associated Staphylococcus aureus MSSA476. The predicted size for the gene, sas0397, is 1503 nucleotides long and the protein, SAS0397, is 500 amino acids. SAS0397 has a mass of 56.21 kDa. Cloning of the gene was performed using the Gateway Technology provided by Invitrogen. Results from SDS-PAGE showed that cloning was successful based on gene expression with a distinct band appearing around 66 kDa in the induced sample. Protein purification will be performed using his-tag column chromatography. Based on bioinformatics, SAS0397 is predicted to be an immunoglobulin G binding protein. Further research to determine its role in the infection pathway is planned. Determining the interaction of SAS0397 during the immune response with B-cells, T-cells, and also its response to normal flora will help to better understand CA-MRSA and provide further information on its invasiveness mechanisms.