PHLIP peptide interaction with a membrane monitored by SAXS

pH (Low) Insertion Peptides (pHLIP peptides) find application in studies of membrane-associated folding because spontaneous insertion of these peptides is conveniently triggered by varying pH. Here, we employed small-angle X-ray scattering (SAXS) to investigate a wild-type (WT) pHLIP peptide oligomeric state in solution at high concentrations and monitor changes in the liposome structure upon peptide insertion into the bilayer. We established that even at high concentrations (up to 300 μM) the WT pHLIP peptide at pH 8.0 does not form oligomers larger than tetramers (which exhibit concentration-dependent transfer to the monomeric state, as was shown previously). This finding has significance for medical applications when high concentration of the peptide is injected into blood and diluted in blood circulation. The interaction of WT pHLIP peptide with liposomes does not alter the unilamellar vesicle structure upon peptide adsorption by the lipid bilayer at high pH or upon insertion across the bilayer at low pH. At the same time, SAXS data clearly demonstrate the insertion of the peptide into the membrane at low pH, which opens the possibility of investigating the kinetic process of polypeptide insertion and exit from the membrane in real time by time-resolved SAXS.