Consequences of a sortase A mutation in Streptococcus gordonii.
Sortase A (SrtA) is required for cell-wall anchoring of LPXTG-containing Gram-positive surface proteins. It was hypothesized, therefore, that disruption of the srtA gene would alter surface anchoring and functions of target LPXTG motif-bearing SspA and SspB proteins of Streptococcus gordonii. Mutant strains in srtA (V288srtA(-), DL1srtA(-)) were constructed in S. gordonii V288 (wtV288) and DL1 (wtDL1). When compared to wtV288, the V288srtA(-) mutant showed decreased biofilm formation on polystyrene, and reduced binding to immobilized purified salivary agglutinin (BIAcore analysis). The wtV288 and V288srtA(-) strains were similar in ultrastructure, but immunogold-labelled SspA/SspB surface expression was reduced on the V288srtA(-) mutant. DL1srtA(-) was also complemented to obtain DL1srtA(+). From the wild-type strains (wtV288, wtDL1), srtA(-) mutants (V288srtA(-), DL1srtA(-)), and the complemented mutant (DL1srtA(+)), cytoplasmic, cell-wall and released extracellular protein fractions were isolated. Each fraction was analysed by SDS-PAGE and immunoblotting with anti-P1. Spent medium from srtA(-) mutant cells contained over-represented proteins, including SspA/SspB (P1 antigen). Mutants showed less P1 on the cell surface than wild-types, as estimated using whole-cell ELISA, and no P1 appeared in the cytoplasmic fractions. Expression of several adhesin genes (sspA/B, cshA/B, fbpA) was generally upregulated in the mutants (V288srtA(-), DL1srtA(-)), but restored to wild-type levels in DL1srtA(+). These data therefore imply that in addition to its role in processing LPXTG-containing adhesins, sortase A has the novel function of contributing to transcriptional regulation of adhesin gene expression.
Department of Diagnostic and Biological Sciences, School of Dentistry, Medical School, University of Minnesota, Minneapolis, MN 55455, USA.
Angela H
AH
Nobbs
Department of Diagnostic and Biological Sciences, School of Dentistry, Medical School, University of Minnesota, Minneapolis, MN 55455, USA.
Reka M
RM
Vajna
Department of Diagnostic and Biological Sciences, School of Dentistry, Medical School, University of Minnesota, Minneapolis, MN 55455, USA.
Jeremy R
JR
Johnson
Department of Diagnostic and Biological Sciences, School of Dentistry, Medical School, University of Minnesota, Minneapolis, MN 55455, USA.
Yongshu
Y
Zhang
Department of Genetics, Cell Biology and Development, Medical School, University of Minnesota, Minneapolis, MN 55455, USA.
Stanley L
SL
Erlandsen
Department of Oral Biology, Joint Health Science Center, University of Florida, Gainesville, FL 32611, USA.
Monika W
MW
Oli
Department of Diagnostic and Biological Sciences, School of Dentistry, Medical School, University of Minnesota, Minneapolis, MN 55455, USA.
Jens
J
Kreth
Department of Oral Biology, Joint Health Science Center, University of Florida, Gainesville, FL 32611, USA.
L Jeannine
LJ
Brady
Mucosal and Vaccine Research Center, Minneapolis VA Medical Center, Minneapolis, MN 55417, USA.|Department of Diagnostic and Biological Sciences, School of Dentistry, Medical School, University of Minnesota, Minneapolis, MN 55455, USA.
Mark C
MC
Herzberg
D018829:Adhesins, Bacterial / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000373:Agglutinins / Q000378:metabolism; D020816:Amino Acid Motifs; D019881:Aminoacyltransferases / Q000235:genetics* / Q000378:metabolism; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D018441:Biofilms / Q000254:growth & development; D003546:Cysteine Endopeptidases / Q000235:genetics* / Q000378:metabolism; D015964:Gene Expression Regulation, Bacterial; D006801:Humans; D008855:Microscopy, Electron, Scanning; D046529:Microscopy, Electron, Transmission; D009154:Mutation; D012463:Saliva / Q000737:chemistry; D054773:Streptococcus gordonii / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development / Q000648:ultrastructure
D018829:Adhesins, Bacterial; D000373:Agglutinins; D001426:Bacterial Proteins; C082314:SspA protein, bacteria; C068465:salivary agglutinin receptor, Streptocococcus; D019881:Aminoacyltransferases; C120542:sortase A; D003546:Cysteine Endopeptidases