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Co‐translational insertion and topogenesis of bacterial membrane proteins monitored in real time
Author(s) -
Mercier Evan,
Wintermeyer Wolfgang,
Rodnina Marina V
Publication year - 2020
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.15252/embj.2019104054
Subject(s) - biology , membrane protein , bacterial protein , membrane , bacteria , microbiology and biotechnology , genetics
Integral membrane proteins insert into the bacterial inner membrane co‐translationally via the translocon. Transmembrane ( TM ) segments of nascent proteins adopt their native topological arrangement with the N‐terminus of the first TM ( TM 1) oriented to the outside (type I) or the inside (type II ) of the cell. Here, we study TM 1 topogenesis during ongoing translation in a bacterial in vitro system, applying real‐time FRET and protease protection assays. We find that TM 1 of the type I protein LepB reaches the translocon immediately upon emerging from the ribosome. In contrast, the type II protein EmrD requires a longer nascent chain before TM 1 reaches the translocon and adopts its topology by looping inside the ribosomal peptide exit tunnel. Looping presumably is mediated by interactions between positive charges at the N‐terminus of TM 1 and negative charges in the tunnel wall. Early TM 1 inversion is abrogated by charge reversal at the N‐terminus. Kinetic analysis also shows that co‐translational membrane insertion of TM 1 is intrinsically rapid and rate‐limited by translation. Thus, the ribosome has an important role in membrane protein topogenesis.