Abstract
The human pathogen Streptococcus pyogenes possesses a chromosomal region, the mga regulon, that contains co-regulated genes important to the virulence of these bacteria. A novel gene located in the mga regulon of a S. pyogenes strain of serotype M1 was cloned and sequenced. It translates into a protein of 305 amino acid residues, including a signal sequence of 32 amino acids and a central region consisting of three tandem repeats. The sequence represents a novel structure with no significant homology to any previously published sequence. The protein was purified from the streptococcal culture media where it is present in substantial amounts. Affinity chromatography of human plasma on Sepharose coupled with the protein specifically absorbed two plasma proteins which were identified as clusterin and histidine-rich glycoprotein (HRG). The interactions between the streptococcal protein and the plasma proteins were further characterized using purified clusterin and HRG. Inhibition experiments indicated that they have affinity for overlapping or closely located sites in the streptococcal protein. Both clusterin and HRG are regulators of the membrane attack complex (C5b-C9) of complement. When the streptococcal protein was added to serum, complement-mediated lysis of sensitized sheep erythrocytes and guinea pig erythrocytes was inhibited. In addition, the streptococcal protein was incorporated into C5b-C9 in serum, indicating the location of its action. The name, protein SIC, streptococcal inhibitor of complement-mediated lysis, is therefore suggested for this novel protein. The occurrence of protein SIC and its gene was investigated in a collection of S. pyogenes strains comprising 55 different M serotypes. Only M1 and M57 strains were positive in this screening, indicating that protein SIC could be a virulence determinant. Thus, during recent years, the M1 serotype has been connected with a world-wide increase of severe and toxic S. pyogenes infections.
Highlights
Streptococcus pyogenes is an important human pathogen causing a number of acute suppurative infections such as ery
The PCR performed with the S3 primer from the 3Ј end of sic and the C1 primer corresponding to a part of the signal sequence of the C5a peptidase gene generated a 2.2-kbp product
It is noteworthy that in contrast to all other described products of the mga regulon, protein SIC does not have the typical structural features of cell wall proteins in Gram-positive bacteria; i.e. a COOH-terminal region anchored to the cell wall through an LPXTG motif (Fischetti et al, 1990; Schneewind et al, 1992, 1995), followed further toward the COOH terminus by a hydrophobic membrane-spanning domain and a tail of mostly positively charged amino acid residues
Summary
Bacterial Strains, Bacteriophage, and Plasmids—Protein SIC and the sic gene were isolated from S. pyogenes strain AP1 (40/58) of the M1 serotype from the Institute of Hygiene and Epidemiology, Prague, Czech Republic. Forty-eight additional strains of different M serotypes were obtained from this institute. These strains have been described (Ben Nasr et al, 1995). S. pyogenes strains of serotypes M3, M33, M42, M52, M61, and M67 were kindly provided by Dr U. Lund University, and a collection of 35 M1 strains isolated in Sweden 1980 – 89 were kindly provided by Dr Stig Holm, Umeå University. Plasmid vectors pUC18/19 (Yanisch-Perron et al, 1985) and pK18/19 (Pridmore, 1987) were used in subcloning experiments. PCR1 products were cloned into the inducible secretion vector pHD389 (Dalboge et al, 1989).
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