Abstract
Bacterial cells utilize monitoring substrates, which undergo force-sensitive translation elongation arrest, to feedback-regulate a Sec-related gene. Vibrio alginolyticus VemP controls the expression of SecD/F that stimulates a late step of translocation by undergoing export-regulated elongation arrest. Here, we attempted at delineating the pathway of the VemP nascent-chain interaction with Sec-related factors, and identified the signal recognition particle (SRP) and PpiD (a membrane-anchored periplasmic chaperone) in addition to other translocon components and a ribosomal protein as interacting partners. Our results showed that SRP is required for the membrane-targeting of VemP, whereas PpiD acts cooperatively with SecD/F in the translocation and arrest-cancelation of VemP. We also identified the conserved Arg-85 residue of VemP as a crucial element that confers PpiD-dependence to VemP and plays an essential role in the regulated arrest-cancelation. We propose a scheme of the arrest-cancelation processes of VemP, which likely monitors late steps in the protein translocation pathway.
Highlights
In bacteria, the evolutionarily conserved Sec translocon and SecA play essential roles in protein translocation across and integration into the membrane (Rapoport et al, 2017)
To identify proteins interacting with a VemP nascent polypeptide, we conducted a systematic PiXie analysis by expressing in E. coli a series of VemP3xFLAG-Myc (VemP-F3M) derivative having pBPA at various positions
VemP was initially produced as the arrested forms, which were converted to full-length mature protein (FL-m) with a half-life of ~0.5 min (Mori et al, 2018)
Summary
The evolutionarily conserved Sec translocon and SecA play essential roles in protein translocation across and integration into the membrane (Rapoport et al, 2017). Bacterial protein secretion to the periplasmic space is thought to occur post-translationally in many cases, some secretory proteins use the signal recognition particle (SRP) pathway, which targets the nascent precursor protein to the translocon (Huber et al, 2005; Shimohata et al, 2005). Even in the latter cases, translocation requires the SecA motor function (Shimohata et al, 2005).
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