Cotranslational recruitment of ribosomes in protocells recreates a translocon-independent mechanism of proteorhodopsin biogenesis

The emergence of lipid membranes and embedded proteins was essential for the evolution of cells. Translocon complexesmediate cotranslational recruitment and membrane insertion of nascent proteins, but they already contain membrane-integral proteins. Therefore, a simpler mechanism must exist, enabling spontaneous membrane integration while preventing aggregation of unchaperoned protein in the aqueous phase. Here, we used giant unilamellar vesicles encapsulating minimal translation components to systematically interrogate the requirements for insertion of the model protein proteorhodopsin (PR) - a structurally ubiquitousmembrane protein. We show that the N-terminal hydrophobic domain of PR is both necessary and sufficient for cotranslational recruitment of ribosomes to the membrane and subsequent membrane insertion of PR. Insertion of N-terminally truncated PR was restored by artificially attaching ribosomes to the membrane. Our findings offer a self-sufficient protein-inherent mechanism as a possible explanation for effective membrane protein biogenesis in a pre-translocon'' era, and they offer new opportunities for generating artificial cells.

Automated summary

The N-terminal hydrophobic domain of the model protein proteorhodopsin (PR) plays a critical role in recruiting ribosomes to the membrane and facilitating its insertion, offering a possible explanation for efficient membrane protein biogenesis in a pre-translocon era. Additionally, artificially attaching ribosomes to the membrane can restore the insertion of N-terminally truncated PR.