Energetics of side-chain snorkeling in transmembrane helices probed by nonproteinogenic amino acids

Cotranslational translocon-mediated insertion of membrane proteins into the endoplasmic reticulum is a key process in membrane protein biogenesis. Although the mechanism is understood in outline, quantitative data on the energetics of the process is scarce. Here, we have measured the effect on membrane integration efficiency of nonproteinogenic analogs of the positively charged amino acids arginine and lysine incorporated into model transmembrane segments. We provide estimates of the influence on the apparent free energy of membrane integration ( ΔG app ) of “snorkeling” of charged amino acids toward the lipid–water interface, and of charge neutralization. We further determine the effect of fluorine atoms and backbone hydrogen bonds (H-bonds) on ΔG app . These results help establish a quantitative basis for our understanding of membrane protein assembly in eukaryotic cells.