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Published on: October 23, 2017
Consequences of a sortase A mutation in Streptococcus gordonii
Angela H Nobbs1, Reka M Vajna1, Jeremy R Johnson1
1Department of Diagnostic and Biological Sciences, School of Dentistry, Medical School, University of Minnesota, Minneapolis, MN 55455, USA.
Microbiology (Reading, England)
|December 1, 2007
Summary
Disrupting Streptococcus gordonii sortase A (SrtA) impacts surface protein anchoring and reduces biofilm formation. SrtA also plays a novel role in regulating adhesin gene expression.
Area of Science:
- Microbiology
- Molecular Biology
- Biochemistry
Background:
- Sortase A (SrtA) is crucial for anchoring LPXTG-motif proteins to the cell wall in Gram-positive bacteria.
- Streptococcus gordonii utilizes SrtA for surface protein display, influencing its interactions and biofilm formation.
Purpose of the Study:
- To investigate the role of sortase A in Streptococcus gordonii by disrupting the srtA gene.
- To determine the impact of srtA disruption on the surface anchoring and function of LPXTG-motif proteins SspA and SspB.
- To explore potential novel functions of SrtA beyond protein anchoring.
Main Methods:
- Construction of srtA mutant strains (V288srtA(-), DL1srtA(-)) in Streptococcus gordonii.
- Assessment of biofilm formation and salivary agglutinin binding using BIAcore analysis.
- Immunogold labeling and whole-cell ELISA to quantify surface protein expression (SspA/SspB, P1 antigen).
- Analysis of protein fractions (cytoplasmic, cell-wall, extracellular) via SDS-PAGE and immunoblotting.
- Gene expression analysis of adhesins (sspA/B, cshA/B, fbpA) in wild-type, mutant, and complemented strains.
Main Results:
- srtA mutants exhibited significantly decreased biofilm formation and reduced binding to salivary agglutinin compared to wild-type strains.
- Surface expression of SspA/SspB and the P1 antigen was reduced in srtA mutants, with no P1 detected in cytoplasmic fractions.
- Spent medium from srtA mutants showed an over-representation of extracellular proteins, including SspA/SspB.
- Expression of several adhesin genes was upregulated in srtA mutants but restored to wild-type levels in the complemented strain.
Conclusions:
- Sortase A is essential for proper cell-wall anchoring of LPXTG-containing adhesins like SspA and SspB in Streptococcus gordonii.
- Disruption of srtA impairs biofilm formation and reduces adherence to salivary components.
- Sortase A possesses a novel function in the transcriptional regulation of adhesin gene expression, in addition to its established role in protein processing and anchoring.
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