Structural characterization of the complex of SecB and metallothionein-labeled proOmpA by cryo-electron microscopy.
ProOmpA is a preprotein that is translocated across the plasma membrane by the general secretory pathway in Escherichia coli. The molecular chaperon SecB in Sec pathway can recognize and bind proOmpA for its translocation. However, the structure of the SecB/proOmpA complex remains unknown. Here, we constructed an uncleavable proOmpA fused with metallothionein at its C-terminus and labeled it with metals in vitro for the study of cryo-electron microscopy. Using single particle cryo-electron microscopy, we reconstructed 3D structure of the stable SecB/proOmpA complex. The structure shows that the major portion of preprotein locates on one side of SecB tetramer, resulting in an asymmetric binding pattern. This work also provides a possible approach to the structure determination of small protein complexes by cryo-electron microscopy.
State Key Laboratory of Biomembrane and Membrane Biotechnology, Center for Structural Biology, School of Life Sciences, Tsinghua University, Beijing, China.
Qiang
Q
Zhou
Shan
S
Sun
Phang
P
Tai
Sen-Fang
SF
Sui
D001425:Bacterial Outer Membrane Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000378:metabolism*; D003001:Cloning, Molecular; D020285:Cryoelectron Microscopy; D029968:Escherichia coli Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D021621:Imaging, Three-Dimensional; D008668:Metallothionein / Q000378:metabolism*; D008670:Metals / Q000378:metabolism; D062105:Molecular Docking Simulation; D011485:Protein Binding; D011498:Protein Precursors / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
D001425:Bacterial Outer Membrane Proteins; D001426:Bacterial Proteins; D029968:Escherichia coli Proteins; D008670:Metals; D011498:Protein Precursors; D011993:Recombinant Fusion Proteins; C057842:SecB protein, Bacteria; C057184:outer membrane protein A precursor (E coli); D008668:Metallothionein