Posttranslational insertion of small membrane proteins by the bacterial signal recognition particle.

Ruth Steinberg,Martin Helmstädter,Ana Natriashvili,Claudine Kraft,Pinku Sarmah,Joen Luirink,Maximilian H Ulbrich,Wei Q Shi,Mariya Licheva,Andrea Origi,Hans-Georg Koch,Stephen High,Princess M Walker,Frederick M Hughson

doi:10.1371/journal.pbio.3000874

Abstract

Small membrane proteins represent a largely unexplored yet abundant class of proteins in pro- and eukaryotes. They essentially consist of a single transmembrane domain and are associated with stress response mechanisms in bacteria. How these proteins are inserted into the bacterial membrane is unknown. Our study revealed that in Escherichia coli, the 27-amino-acid-long model protein YohP is recognized by the signal recognition particle (SRP), as indicated by in vivo and in vitro site-directed cross-linking. Cross-links to SRP were also observed for a second small membrane protein, the 33-amino-acid-long YkgR. However, in contrast to the canonical cotranslational recognition by SRP, SRP was found to bind to YohP posttranslationally. In vitro protein transport assays in the presence of a SecY inhibitor and proteoliposome studies demonstrated that SRP and its receptor FtsY are essential for the posttranslational membrane insertion of YohP by either the SecYEG translocon or by the YidC insertase. Furthermore, our data showed that the yohP mRNA localized preferentially and translation-independently to the bacterial membrane in vivo. In summary, our data revealed that YohP engages an unique SRP-dependent posttranslational insertion pathway that is likely preceded by an mRNA targeting step. This further highlights the enormous plasticity of bacterial protein transport machineries.

Highlights

The punctate appearance of YohP-green fluorescent protein (GFP) in vivo could result from aggregation or from internal membranes, which bacteria occasionally produce upon membrane protein expression [34]
We have identified an unexpected signal recognition particle (SRP)-dependent posttranslational insertion mode for the small membrane protein YohP
One example is YohP, which was studied here and which is highly enriched in the inner membrane of E. coli (Fig 1)

Summary

Introduction

We analyzed the targeting and insertion mode of the small membrane protein YohP bearing no or only short C-terminal tags and identified a novel SRP-dependent posttranslational protein insertion pathway in bacteria. In order to establish whether SRP is involved in the insertion of other small membrane proteins, the in vitro cross-linking experiment was performed with YkgR (Fig 1A).

Results

Conclusion