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Co‐translational insertion and topogenesis of bacterial membrane proteins monitored in real time

MPS-Authors
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Mercier,  E.
Department of Physical Biochemistry, MPI for Biophysical Chemistry, Max Planck Society;

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Wintermeyer,  W.
Research Group of Ribosome Dynamics, MPI for biophysical chemistry, Max Planck Society;

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Rodnina,  M. V.
Department of Physical Biochemistry, MPI for Biophysical Chemistry, Max Planck Society;

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Citation

Mercier, E., Wintermeyer, W., & Rodnina, M. V. (2020). Co‐translational insertion and topogenesis of bacterial membrane proteins monitored in real time. EMBO Journal, 39(15): e104054. doi:10.15252/embj.2019104054.


Cite as: http://hdl.handle.net/21.11116/0000-0006-8AB0-7
Abstract
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.