Roles of sortase in surface expression of the major protein adhesin P1, saliva-induced aggregation and adherence, and cariogenicity of Streptococcus mutans.
Sortase is a newly discovered transpeptidase that covalently links LPXTGX-containing surface proteins to the gram-positive bacterial cell wall. In this study, the sortase gene (srtA) was isolated from Streptococcus mutans NG8 by PCR. The gene encoded a 246-amino-acid protein, including a 40-amino-acid signal peptide. The srtA gene was insertionally inactivated by a tetracycline resistance cassette. P1, a major surface protein adhesin previously shown to anchor to the peptidoglycan by the LPXTGX motif, was secreted into the culture medium by the srtA mutant. In contrast, the wild-type P1 remained cell wall associated. Complementation of the mutant with srtA restored the P1 surface expression phenotype. P1 produced by the mutant, but not that produced by the wild type and the srtA-complemented mutant, was recognized by an antibody raised against the hydrophobic domain and charged tail C terminal to the LPXTGX motif. These results suggest that the failure to anchor P1 to the cell wall is due to the lack of cleavage of P1 at the LPXTGX motif. The srtA mutant was markedly less hydrophobic than the wild type and the complemented mutant. The srtA mutant failed to aggregate in the presence of saliva or salivary agglutinin and adhered poorly to saliva- or salivary agglutinin-coated hydroxylapatite. In rats, the srtA mutant colonized the teeth poorly when sucrose was absent. When sucrose was present, the srtA mutant colonized the teeth but less effectively and induced significantly less caries (P < 0.05) than the wild-type strain. In conclusion, the sortase enzyme in S. mutans is responsible for anchoring P1 to the cell surface and plays a role in modulating the surface properties and cariogenicity of S. mutans.
Lee
Song F
SF
Department of Applied Oral Sciences, Faculty of Dentistry, Dalhousie University, Halifax, Nova Scotia, Canada B3H 3J5. Song.Lee@Dal.Ca
Boran
Thomas L
TL
D019881:Aminoacyltransferases / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001422:Bacterial Adhesion; D001426:Bacterial Proteins / Q000378:metabolism*; D003546:Cysteine Endopeptidases; D003731:Dental Caries / Q000382:microbiology* / Q000503:physiopathology; D005260:Female; D005816:Genetic Complementation Test; D008562:Membrane Glycoproteins; D008969:Molecular Sequence Data; D009061:Mouth Mucosa / Q000382:microbiology; D009154:Mutation; D051381:Rats; D017207:Rats, Sprague-Dawley; D012463:Saliva / Q000382:microbiology* / Q000502:physiology; D013290:Streptococcal Infections / Q000382:microbiology / Q000503:physiopathology; D013295:Streptococcus mutans / Q000201:enzymology / Q000472:pathogenicity* / Q000502:physiology; D014070:Tooth / Q000382:microbiology; D014774:Virulence
D001426:Bacterial Proteins; D008562:Membrane Glycoproteins; C055764:S-layer proteins; D019881:Aminoacyltransferases; C120542:sortase A; D003546:Cysteine Endopeptidases