Abstract：
Electrogenic bacteria deliver excess respiratory electrons to externally located metal oxide particles and electrodes. The biochemical basis for this process is arguably best understood for species of Shewanella where the integral membrane complex termed MtrCAB is key to electron transfer across the bacterial outer membranes. A crystal structure was recently resolved for MtrCAB from S. baltica OS185. However, X-ray diffraction did not resolve the N-terminal residues so that the lipidation status of proteins in the mature complex was poorly described. Here we report liquid chromatography mass spectrometry revealing the intact mass values for all three proteins in the MtrCAB complexes purified from Shewanella oneidensis MR-1 and S. baltica OS185. The masses of MtrA and MtrB are consistent with both proteins being processed by Signal Peptidase I and covalent attachment of ten c-type hemes to MtrA. The mass of MtrC is most reasonably interpreted as arising from protein processed by Signal Peptidase II to produce a diacylated lipoprotein containing ten c-type hemes. Our two-step protocol for liquid-chromatography mass spectrometry used a reverse phase column to achieve on-column detergent removal prior to gradient protein resolution and elution. We envisage the method will be capable of simultaneously resolving the intact mass values for multiple proteins in other membrane protein complexes.
摘要：
产电细菌将过量的呼吸电子递送到位于外部的金属氧化物颗粒和电极。可以说，对于希瓦氏菌属物种，可以最好地理解该过程的生化基础，其中称为MtrCAB的完整膜复合物是跨细菌外膜电子转移的关键。最近从S.balticaOS185解析了MtrCAB的晶体结构。然而,X射线衍射不能分解N末端残基，因此成熟复合物中蛋白质的脂化状态描述不佳。在这里，我们报告了液相色谱质谱，揭示了从ShewanellaoneidensisMR-1和S.balticaOS185中纯化的MtrCAB复合物中所有三种蛋白质的完整质量值。MtrA和MtrB的质量与信号肽酶I和十个c型血红素与MtrA的共价连接所处理的两种蛋白质一致。MtrC的质量被最合理地解释为由信号肽酶II加工的蛋白质产生，以产生包含十个c型血红素的二酰化脂蛋白。我们的液相色谱质谱两步方案使用反相柱在梯度蛋白质分离和洗脱之前实现柱上去污剂去除。我们设想该方法将能够同时解析其他膜蛋白复合物中多种蛋白的完整质量值。
