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.
Nobbs
Angela H
AH
Department of Diagnostic and Biological Sciences, School of Dentistry, Medical School, University of Minnesota, Minneapolis, MN 55455, USA.
Vajna
Reka M
RM
Department of Diagnostic and Biological Sciences, School of Dentistry, Medical School, University of Minnesota, Minneapolis, MN 55455, USA.
Johnson
Jeremy R
JR
Department of Diagnostic and Biological Sciences, School of Dentistry, Medical School, University of Minnesota, Minneapolis, MN 55455, USA.
Zhang
Yongshu
Y
Department of Diagnostic and Biological Sciences, School of Dentistry, Medical School, University of Minnesota, Minneapolis, MN 55455, USA.
Erlandsen
Stanley L
SL
Department of Genetics, Cell Biology and Development, Medical School, University of Minnesota, Minneapolis, MN 55455, USA.
Oli
Monika W
MW
Department of Oral Biology, Joint Health Science Center, University of Florida, Gainesville, FL 32611, USA.
Kreth
Jens
J
Department of Diagnostic and Biological Sciences, School of Dentistry, Medical School, University of Minnesota, Minneapolis, MN 55455, USA.
Brady
L Jeannine
LJ
Department of Oral Biology, Joint Health Science Center, University of Florida, Gainesville, FL 32611, USA.
Herzberg
Mark C
MC
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.
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