Solid State NMR Studies on Acylated Transmembrane Peptides Driving Membrane Fusion

Abstract

The fusion of biological membranes is mediated by integral membrane proteins with α-helical transmembrane segments (TMSs). Additionally, those proteins are often modified by the covalent attachment of hydrocarbon chains. Previously, a series of de novo designed α-helical peptides with mixed Leu/Val sequences was presented, mimicking fusogenic TMSs in model membranes (Hofmann et al., Proc. Natl. Acad. Sci. USA 101 (2004) 14776-14781). From this series, we have investigated the peptide LV16 (KKKWLVLVLVLVLVLVLVLVKKK), which was synthesized presenting either a free N-terminus or an N-acylation of 2, 8, 12, or 16 carbons. We used 2H and 31P NMR spectroscopy to investigate the structure and dynamics of those peptide lipid modifications in POPC and DLPC bilayers and compared them to the hydrocarbon chains of the surrounding membrane. Except for the C-2 chain, all peptide acyl chains were found to insert well into the membrane. This can be understood from the high local lipid concentration, which the N-terminal lipid chains experience. The insertion of these peptides did not influence the membrane structure and dynamics as seen from 2H and 31P NMR. In spite of the fact that the longer acyl chains insert into the membrane, there is no perfect length adaptation. Even the C-16 chain on the peptide, which could match the length of the POPC palmitoyl chain exhibited lower order parameters in the upper chain, which get closer and finally reach similar values in the lower chain region. 2H NMR square law plots reveal a slightly more dynamic characteristic of the peptide acyl chains compared to the surrounding phospholipids. In spite of the significantly different chain lengths of the acyl chains, the fraction of gauche defects in the inserted chains is constant, suggesting similar entropies of the inserted chains.