Membrane-associated folding and unfolding

We are studying the molecular events that occur when a peptide inserts across a membrane or exits from it. Using pH jumps to trigger insertion/exit of the pHLIP (pH Low Insertion Peptide) to enable kinetic analysis, we show that insertion occurs in several steps, with rapid (0.1 sec) interfacial helix formation followed by a much slower (100 sec) insertion pathway to form a transmembrane helix. The reverse process of unfolding and peptide exit from the bilayer core, which can be induced by a rapid pH jump from acidic to basic, proceeds much faster than folding/insertion and through different intermediate states. In the exit pathway, the helix-coil transition is initiated while the polypeptide is still inside the membrane. We also designed two pHLIP-variants where Asp and Glu residues were removed from the C-terminus, which inserts across the membrane. The variants preserve the same pH-dependent properties of pHLIP peptide interaction with the membrane, but insertion occurs 10-30 times faster than in the case of the parent pHLIP peptide. A kinetic model of peptide-membrane insertion/folding and exit/unfolding will be discussed. The work was supported by grant from the NIH RO1133890 to OAA, DME, YRK.