Abstract

The heterotrimeric SecYEG complex comprises a protein-conducting channel in the bacterial cytoplasmic membrane. SecYEG functions together with the motor protein SecA in preprotein translocation. Here, we have addressed the functional oligomeric state of SecYEG when actively engaged in preprotein translocation. We reconstituted functional SecYEG complexes labelled with fluorescent markers into giant unilamellar vesicles at a natively low density. Förster's resonance energy transfer and fluorescence (cross-) correlation spectroscopy with single-molecule sensitivity allowed for independent observations of the SecYEG and preprotein dynamics, as well as complex formation. In the presence of ATP and SecA up to 80% of the SecYEG complexes were loaded with a preprotein translocation intermediate. Neither the interaction with SecA nor preprotein translocation resulted in the formation of SecYEG oligomers, whereas such oligomers can be detected when enforced by crosslinking. These data imply that the SecYEG monomer is sufficient to form a functional translocon in the lipid membrane.

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Authors

Kedrov A, Kusters I, Krasnikov VV, Driessen AJ

Institution

Department of Molecular Microbiology, Groningen Biomolecular Science and Biotechnology Institute, The Zernike Institute for Advanced Materials, University of Groningen, Groningen, The Netherlands.

Source

The EMBO journal 30:21 2011 Nov 2 pg 4387-97

MeSH

Escherichia coli
Escherichia coli Proteins
Gene Dosage
Membrane Lipids
Membrane Proteins
Methanococcus
Models, Biological
Organisms, Genetically Modified
Protein Multimerization
Protein Precursors
Protein Transport
Unilamellar Liposomes

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

21897368