Proline affects oligomerization of a coiled coil by inducing a kink in a long helix

The structural effect of a proline in a helix in trifluoroethanol (TFE)/wat er medium was examined on a 29-mer peptide and its proline analog derived f rom the leucine zipper (LZ)-like motif of gp41 (the transmembrane glycoprot ein of HIV-1) by NMR and circular dichroism (CD) spectroscopies. Lower heli cal content was found for the proline mutant from the CD study. NMR data sh ow that distortion of the helix by proline is local and occurs mainly on th e N-terminal side of the substitution site. Molecular dynamics computation exhibits a bending of the helical axis of 30 degrees +/- 10 degrees, in agr eement with X-ray diffraction results. Light-scattering experiments indicat ed that the average aggregation number of the proline-substituted mutant is substantially lower than that of the wild-type peptide. From the ratio of dissociation constants of the wild-type and the proline mutant peptides, th e difference in free energy of trimeric formation is calculated to be 2.1 k cal/mol, Thermal stability, helicity, and the average aggregation number fo r the helix oligomers were found to be correlated. The structural alteratio n and the reduced coiled coil stability may account for the deficiency in t he biological functions of the proline mutants of gp41 and in the inhibitor y action of proline-substituted peptides, These effects may also be importa nt in unraveling the roles played by proline in transmembrane proteins. (C) 1999 Academic Press.