Department of Medical Microbiology, University of Groningen and University Medical Center Groningen, Groningen, The Netherlands.
Proteomics. 2012 Oct;12(19-20):3049-62. doi: 10.1002/pmic.201200144. Epub 2012 Sep 20.
Sortases catalyze the covalent attachment of proteins with a C-terminal LPxTG motif to the cell walls of Gram-positive bacteria. Here, we show that deletion of the srtA genes of Staphylococcus aureus and Staphylococcus epidermidis resulted in the dislocation of several LPxTG proteins from the cell wall to the growth medium. Nevertheless, proteomics and Western blotting analyses revealed that substantial amounts of the identified proteins remained cell wall bound through noncovalent interactions. The protein dislocation phenotypes of srtA mutants of S. aureus and S. epidermidis were reverted by ectopic expression of srtA genes of either species. Interestingly, S. epidermidis contains a second sortase A, which was previously annotated as ``SrtC.'' Ectopic expression of this SrtC in srtA mutant cells reverted the dislocation of some, but not all, cell wall associated proteins. Similarly, defects in biofilm formation were reverted by ectopic expression of SrtC in some, but not all, tested srtA mutant strains. Finally, overexpression of SrtA resulted in increased levels of biofilm formation in some tested strains. Taken together, these findings show that the substrate specificities of SrtA and SrtC overlap partially, and that sortase levels may be limiting for biofilm formation in some staphylococci.
Sortases 催化具有 C 末端 LPxTG 基序的蛋白质与革兰氏阳性菌细胞壁的共价连接。在这里，我们表明，金黄色葡萄球菌和表皮葡萄球菌的 srtA 基因缺失导致几种 LPxTG 蛋白从细胞壁脱位到生长培养基中。然而，蛋白质组学和 Western blot 分析表明，通过非共价相互作用，大量鉴定的蛋白质仍然与细胞壁结合。金黄色葡萄球菌和表皮葡萄球菌的 srtA 突变体的蛋白质脱位表型通过同种或异种 srtA 基因的异位表达得到逆转。有趣的是，表皮葡萄球菌含有第二种 Sortase A，以前被注释为“SrtC”。该 SrtC 在 srtA 突变细胞中的异位表达逆转了一些，但不是所有，与细胞壁相关的蛋白质的脱位。类似地，通过在一些但不是所有测试的 srtA 突变菌株中异位表达 SrtC，生物膜形成缺陷得到了逆转。最后，在一些测试菌株中，SrtA 的过表达导致生物膜形成水平增加。总之，这些发现表明 SrtA 和 SrtC 的底物特异性部分重叠，并且在某些葡萄球菌中，分选酶水平可能限制生物膜形成。
