In vivo identification of the outer membrane protein OmcA-MtrC interaction network in Shewanella oneidensis MR-1 cells using novel hydrophobic chemical cross-linkers.
In vivo identification of the outer membrane protein OmcA-MtrC interaction network in Shewanella oneidensis MR-1 cells using novel hydrophobic chemical cross-linkers.
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DOI：
10.1021/pr7007658
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发表时间：
2008-02
影响因子：
4.4
通讯作者：
Haizhen Zhang;Xiaoting Tang;G. Munske;Natalia L Zakharova;Li Yang;Chunxiang Zheng;M. Wolff;N. Tolić;G. Anderson;Liang Shi;M. Marshall;J. Fredrickson;J. Bruce
中科院分区：
文献类型：
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作者：
Haizhen Zhang;Xiaoting Tang;G. Munske;Natalia L Zakharova;Li Yang;Chunxiang Zheng;M. Wolff;N. Tolić;G. Anderson;Liang Shi;M. Marshall;J. Fredrickson;J. Bruce
外膜（OM）细胞色素OmcA（SO 1779）和MtrC（SO 1778）是希瓦氏菌（Shewanella oneidensis）进行金属（氢）氧化物厌氧呼吸的电子传递的组成部分。在这里，OmcA-MtrC相互作用在体内使用一种新的疏水化学交联剂（MRN）结合免疫沉淀技术进行了鉴定。此外，从交联复合物中鉴定其他OM蛋白允许OmcA-MtrC相互作用网络的首次可视化。对omcA和mtrC突变体细胞的进一步实验表明，OmcA在网络相互作用中起核心作用。为了进行比较，还平行使用了两种商业交联剂，两者均导致较少的OM蛋白鉴定，表明MRN用于鉴定膜蛋白相互作用的上级性质。最后，体内交联和细胞裂解物交联的比较实验导致显著不同的蛋白质相互作用数据，证明了体内交联对于研究细胞中蛋白质-蛋白质相互作用的重要性。
Outer membrane (OM) cytochromes OmcA (SO1779) and MtrC (SO1778) are the integral components of electron transfer used by Shewanella oneidensis for anaerobic respiration of metal (hydr)oxides. Here the OmcA-MtrC interaction was identified in vivo using a novel hydrophobic chemical cross-linker (MRN) combined with immunoprecipitation techniques. In addition, identification of other OM proteins from the cross-linked complexes allows first visualization of the OmcA-MtrC interaction network. Further experiments on omcA and mtrC mutant cells showed OmcA plays a central role in the network interaction. For comparison, two commercial cross-linkers were also used in parallel, and both resulted in fewer OM protein identifications, indicating the superior properties of MRN for identification of membrane protein interactions. Finally, comparison experiments of in vivo cross-linking and cell lysate cross-linking resulted in significantly different protein interaction data, demonstrating the importance of in vivo cross-linking for study of protein-protein interactions in cells.