Department of Structural Infection Biology, Center for Structural Systems Biology (CSSB) & Helmholtz-Center for Infection Research (HZI), Hamburg, Germany.
Division of Biophysics, Research Center Borstel, Leibniz Lung Center, Borstel, Germany.
PLoS Pathog. 2021 Nov 17;17(11):e1010052. doi: 10.1371/journal.ppat.1010052. eCollection 2021 Nov.
The methyltransferase FliB posttranslationally modifies surface-exposed ɛ-N-lysine residues of flagellin, the protomer of the flagellar filament in Salmonella enterica (S. enterica). Flagellin methylation, reported originally in 1959, was recently shown to enhance host cell adhesion and invasion by increasing the flagellar hydrophobicity. The role of FliB in this process, however, remained enigmatic. In this study, we investigated the properties and mechanisms of FliB from S. enterica in vivo and in vitro. We show that FliB is an S-adenosylmethionine (SAM) dependent methyltransferase, forming a membrane associated oligomer that modifies flagellin in the bacterial cytosol. Using X-band electron paramagnetic resonance (EPR) spectroscopy, zero-field 57Fe Mössbauer spectroscopy, methylation assays and chromatography coupled mass spectrometry (MS) analysis, we further found that FliB contains an oxygen sensitive [4Fe-4S] cluster that is essential for the methyl transfer reaction and might mediate a radical mechanism. Our data indicate that the [4Fe-4S] cluster is coordinated by a cysteine rich motif in FliB that is highly conserved among multiple genera of the Enterobacteriaceae family.
甲基转移酶 FliB 对鞭毛蛋白的ɛ-N-赖氨酸残基进行翻译后修饰，鞭毛蛋白是沙门氏菌（S. enterica）鞭毛丝的原体。1959 年首次报道的鞭毛蛋白甲基化最近被证明通过增加鞭毛疏水性来增强宿主细胞的粘附和侵袭。然而，FliB 在这个过程中的作用仍然是个谜。在这项研究中，我们在体内和体外研究了来自沙门氏菌的 FliB 的特性和机制。我们表明 FliB 是一种 S-腺苷甲硫氨酸（SAM）依赖性甲基转移酶，形成一种与膜相关的寡聚体，在细菌细胞质中修饰鞭毛蛋白。使用 X 波段电子顺磁共振（EPR）光谱、零场 57Fe Mössbauer 光谱、甲基化测定和色谱偶联质谱（MS）分析，我们还发现 FliB 含有一个对甲基转移反应至关重要的氧敏感[4Fe-4S]簇，并且可能介导一个自由基机制。我们的数据表明，[4Fe-4S]簇由 FliB 中富含半胱氨酸的基序配位，该基序在肠杆菌科的多个属中高度保守。
