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

As part of a program to study the effect of glycosylation an the three-dimensional structures of HIV-1(IIIB) V3 peptide constructs, we have examined the solution structures of a 15 residue peptide (RIQRGPGRAFVTIGK, P18(IIIB)), originally mapped as an epitope recognized by CD8(+) D-d class I MHC-restricted murine cytotoxic T-lymphocytes (CTL), and an analogue (P18(IIIB)-g), O-glycosylated with an alpha-galactosamine on Thr-12, using NMR, circular dichroism and molecular modeling methods, Our studies show Chat 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 famed around the immunodominant tip (G(5)-R(8)). Glycosylation on T-12 'tightens' the turn slightly as suggested by NOE and CD analysis, Ln 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 mag. 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 P18(IIIB) was completely abrogated by threonine glycosylation.