Structural comparison of a 15 residue peptide from the V3 loop of HIV-1IIIb and an O-glycosylated analogue.

As part of a program to study the effect of glycosylation on the three-dimensional structures of HIV-1IIIB V3 peptide constructs, we have examined the solution structures of a 15 residue peptide (RIQRGPGRAFVTIGK, P18IIIB)- originally mapped as an epitope recognized by CD8+ Dd class I MHC-restricted murine cytotoxic T-lymphocytes (CTL), and an analogue (P18IIIB-g), O-glycosylated with an alpha-galactosamine on Thr-12, using NMR, circular dichroism and molecular modeling methods. Our studies show that the peptides sample mainly random conformations in aqueous solution near 25 degrees C and become more ordered by the addition of trifluoroethanol. Upon decreasing the temperature to 5 degrees C, a reverse turn is formed around the immunodominant tip (G5-R8). Glycosylation on T12 'tightens' the turn slightly as suggested by NOE and CD analysis. In addition, the sugar has a defined conformation with respect to the peptide backbone and influences the local peptide conformation. These data suggest that simple glycosylation may influence the conformational equilibrium of a V3 peptide which contains a domain critical for antibody recognition and virus neutralization. We also show that the ability of cytotoxic T-lymphocytes (CTL) to lyse tumor cells presenting P18IIIB was completely abrogated by threonine glycosylation.