Institute for Microbiology, Ernst-Moritz-Arndt-University of Greifswald, Greifswald, Germany.
Antioxid Redox Signal. 2013 Apr 10;18(11):1273-95. doi: 10.1089/ars.2012.4686. Epub 2012 Oct 18.
Protein S-bacillithiolations are mixed disulfides between protein thiols and the bacillithiol (BSH) redox buffer that occur in response to NaOCl in Bacillus subtilis. We used BSH-specific immunoblots, shotgun liquid chromatography (LC)-tandem mass spectrometry (MS/MS) analysis and redox proteomics to characterize the S-bacillithiolomes of B. subtilis, B. megaterium, B. pumilus, B. amyloliquefaciens, and Staphylococcus carnosus and also measured the BSH/oxidized bacillithiol disulfide (BSSB) redox ratio after NaOCl stress.
In total, 54 proteins with characteristic S-bacillithiolation (SSB) sites were identified, including 29 unique proteins and eight proteins conserved in two or more of these bacteria. The methionine synthase MetE is the most abundant S-bacillithiolated protein in Bacillus species after NaOCl exposure. Further, S-bacillithiolated proteins include the translation elongation factor EF-Tu and aminoacyl-tRNA synthetases (ThrS), the DnaK and GrpE chaperones, the two-Cys peroxiredoxin YkuU, the ferredoxin-NADP(+) oxidoreductase YumC, the inorganic pyrophosphatase PpaC, the inosine-5'-monophosphate dehydrogenase GuaB, proteins involved in thiamine biosynthesis (ThiG and ThiM), queuosine biosynthesis (QueF), biosynthesis of aromatic amino acids (AroA and AroE), serine (SerA), branched-chain amino acids (YwaA), and homocysteine (LuxS and MetI). The thioredoxin-like proteins, YphP and YtxJ, are S-bacillithiolated at their active sites, suggesting a function in the de-bacillithiolation process. S-bacillithiolation is accompanied by a two-fold increase in the BSSB level and a decrease in the BSH/BSSB redox ratio in B. subtilis.
Many essential and conserved proteins, including the dominant MetE, were identified in the S-bacillithiolome of different Bacillus species and S. carnosus using shotgun-LC-MS/MS analyses.
S-bacillithiolation is a widespread redox control mechanism among Firmicutes bacteria that protects conserved metabolic enzymes and essential proteins against overoxidation.
蛋白 S-芽孢硫醇化是蛋白巯基与芽孢硫醇（BSH）氧化还原缓冲剂之间的混合二硫键，发生于枯草芽孢杆菌对次氯酸钠的反应中。我们使用 BSH 特异性免疫印迹、液相色谱（LC）-串联质谱（MS/MS）分析和氧化还原蛋白质组学来描述枯草芽孢杆菌、巨大芽孢杆菌、短小芽孢杆菌、解淀粉芽孢杆菌和肉葡萄球菌的 S-芽孢硫醇组，并在次氯酸钠应激后测量 BSH/氧化芽孢硫醇二硫化物（BSSB）氧化还原比。
共鉴定出 54 种具有特征性 S-芽孢硫醇化（SSB）位点的蛋白，包括 29 种独特蛋白和 8 种在两种或两种以上细菌中保守的蛋白。甲硫氨酸合酶 MetE 是枯草芽孢杆菌在次氯酸钠暴露后最丰富的 S-芽孢硫醇化蛋白。此外，S-芽孢硫醇化蛋白包括翻译延伸因子 EF-Tu 和氨酰-tRNA 合成酶（ThrS）、DnaK 和 GrpE 伴侣蛋白、双-Cys 过氧化物酶 YkuU、铁氧还蛋白-NADP（+）氧化还原酶 YumC、无机焦磷酸酶 PpaC、肌苷-5'-单磷酸脱氢酶 GuaB、参与硫胺素生物合成的蛋白（ThiG 和 ThiM）、Queuosine 生物合成（QueF）、芳香族氨基酸生物合成（AroA 和 AroE）、丝氨酸（SerA）、支链氨基酸（YwaA）和同型半胱氨酸（LuxS 和 MetI）。硫氧还蛋白样蛋白 YphP 和 YtxJ 在其活性位点上被 S-芽孢硫醇化，这表明它们在去芽孢硫醇化过程中具有功能。S-芽孢硫醇化伴随着枯草芽孢杆菌 BSSB 水平增加两倍，BSH/BSSB 氧化还原比降低。
使用液相色谱-MS/MS 分析，在不同的枯草芽孢杆菌和肉葡萄球菌中鉴定到许多必需和保守的蛋白，包括占主导地位的 MetE，这些蛋白都存在于 S-芽孢硫醇组中。
S-芽孢硫醇化是厚壁菌门细菌中广泛存在的氧化还原调控机制，可保护保守的代谢酶和必需蛋白免受过度氧化。
