title: Cross-linking-based flexibility and proximity relationships between the TM segments of the Escherichia coli YidC.
abstract: The YidC family members function to insert proteins into membranes in bacteria, chloroplasts, and mitochondria, and they can also act as a platform to fold and assemble proteins into higher-order complexes. Here, we provide information about the proximity relationships and dynamics of the five conserved C-terminal transmembrane (TM) regions within Escherichia coli YidC. By using a YidC construct with tandem thrombin protease sites introduced into the cytoplasmic loop C1, cross-linking between paired-Cys residues located within TM segments or in the membrane border regions was studied using thio-specific homobifunctional cross-linking agents with different spanner lengths or by iodine-catalyzed disulfide formation. These in vivo cross-linking studies that can detect transient interactions and different conformational states of the protein show that TM3, TM4, TM5, and TM6 each have a face oriented toward TM2 of the in vivo expressed YidC. The studies also reveal that YidC is a dynamic protein, as cross-linking was observed between cytoplasmic Cys residues with a variety of cross-linkers. A large number of conserved proline residues on the cytoplasmic side of the five conserved core TM segments could explain the observed flexibility, and the structural fluctuations of the TM segments could provide an explanation for how YidC is able to recognize a variety of different substrates.
authors.affiliation: Department of Chemistry and Biochemistry, The Ohio State University , Columbus, Ohio 43210, United States.
authors.forename: Seth W
authors.initials: SW
authors.lastname: Hennon
authors.forename: Ross E
authors.initials: RE
authors.lastname: Dalbey
mesh_terms: D002462:Cell Membrane / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D017124:Conserved Sequence; D003432:Cross-Linking Reagents / Q000737:chemistry; D004220:Disulfides / Q000737:chemistry; D004926:Escherichia coli; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011392:Proline / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D017510:Protein Folding; D021381:Protein Transport / Q000502:physiology
chemical_list: D003432:Cross-Linking Reagents; D004220:Disulfides; D029968:Escherichia coli Proteins; D026901:Membrane Transport Proteins; C412443:YIDC protein, E coli; D011392:Proline