1887

Abstract

We investigated culture supernatant proteins from the M1 serotype of by two-dimensional gel electrophoresis and peptide mass mapping analysis, and characterized the single protein spots. Among them, we analysed the Spy0747 protein. This protein is homologous to the SsnA protein, a cell-wall-located DNase expressed in serotype 2. We designated the Spy0747 protein as SpnA. SpnA protein was also detected in the insoluble fraction of whole-cell lysates using shotgun proteomic analysis, suggesting that SpnA is also located in the cell wall. SpnA was expressed as a glutathione -transferase-fusion protein in . We confirmed that the recombinant protein had DNase activity that was dependent on Ca and Mg, like SsnA. Blood bactericidal assays and mouse infection model experiments showed that the knockout strain was less virulent than the parental strain, thus suggesting that SpnA could play an important role in virulence. Using PCR, we found that the gene was present in all clinical strains we examined. Our results, together with a previous report identifying Spy0747 as a surface-associated protein, suggest that SpnA is an important cell-wall-located DNase that is generally produced in and is involved in virulence.

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2010-01-01
2019-11-18
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vol. , part 1, pp. 184 - 190

Comparative proteomic analysis. Proteins were identified in soluble and insoluble fractions of SF370. r, calculated molecular mass; pI, calculated isoelectric point; MSD, number of membrane-spanning domains estimated with SOSUI; SP, the possibility value with SignalP hidden-Markov model. The number in each Soluble and Insoluble column indicates the average number of assigned peptide fragments. [ Excel file] (69 kb)



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