title: In vivo identification of the outer membrane protein OmcA-MtrC interaction network in Shewanella oneidensis MR-1 cells using novel hydrophobic chemical cross-linkers.
abstract: 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.
authors.affiliation: Department of Chemistry, Washington State University, Pullman, Washington, USA.
authors.forename: Haizhen
authors.initials: H
authors.lastname: Zhang
authors.forename: Xiaoting
authors.initials: X
authors.lastname: Tang
authors.forename: Gerhard R
authors.initials: GR
authors.lastname: Munske
authors.forename: Natalia
authors.initials: N
authors.lastname: Zakharova
authors.forename: Li
authors.initials: L
authors.lastname: Yang
authors.forename: Chunxiang
authors.initials: C
authors.lastname: Zheng
authors.forename: Megan A
authors.initials: MA
authors.lastname: Wolff
authors.forename: Nikola
authors.initials: N
authors.lastname: Tolic
authors.forename: Gordon A
authors.initials: GA
authors.lastname: Anderson
authors.forename: Liang
authors.initials: L
authors.lastname: Shi
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Marshall
authors.forename: James K
authors.initials: JK
authors.lastname: Fredrickson
authors.forename: James E
authors.initials: JE
authors.lastname: Bruce
mesh_terms: D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D015153:Blotting, Western; D003432:Cross-Linking Reagents / Q000138:chemical synthesis / Q000737:chemistry*; D003574:Cytochrome c Group / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D017353:Gene Deletion; D057927:Hydrophobic and Hydrophilic Interactions; D047468:Immunoprecipitation; D011485:Protein Binding; D025941:Protein Interaction Mapping; D020592:Shewanella / Q000378:metabolism*; D053719:Tandem Mass Spectrometry
chemical_list: D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D003432:Cross-Linking Reagents; D003574:Cytochrome c Group; C433527:MtrC protein, Shewanella