Proteomic Analysis and Identification of Streptococcus pyogenes Surface-Associated Proteins

Wyeth Vaccine Research, Pearl River, New York and Wyeth Research, Cambridge, Massachusetts

^* To whom correspondence should be addressed. Email: olmsteds{at}wyeth.com.

	Abstract

Streptococcus pyogenes is a gram-positive human pathogen that causes a wide spectrum of disease, placing a significant burden on public health. Bacterial surface-associated proteins play crucial roles in host-pathogen interactions and pathogenesis, and are important targets for the immune system. The identification of these proteins for vaccine development is an important goal of bacterial proteomics. Here we describe a method of proteolytic digestion of surface-exposed proteins to identify surface antigens of S. pyogenes. Peptides generated by trypsin digestion were analyzed by multi-dimensional tandem mass spectrometry. This approach allowed the identification of 79 proteins on the bacterial surface, including 14 proteins containing cell wall-anchoring motifs, 12 lipoproteins, 9 secreted proteins, 22 membrane-associated proteins, one bacteriophage-associated protein, and 21 proteins commonly identified as cytoplasmic. Thirty-three of these proteins have not been previously identified as cell-surface associated in S. pyogenes. Several proteins were expressed in Escherichia coli and the purified proteins used to generate specific mouse antisera for use in a whole-cell enzyme-linked immunosorbent assay. The immunoreactivity of specific antisera to some of these antigens confirmed their surface localization. The data reported here will provide guidance to the development of a novel vaccine to prevent infections caused by S. pyogenes.