Structure of segments of a G protein-coupled receptor: CD and NMR analysis of the Saccharomyces cerevisiae tridecapeptide pheromone receptor.

Peptides representing both loop and the sixth transmembrane regions of the alpha-factor receptor of Saccharomyces cerevisiae were synthesized by solid-phase procedures and purified to near homogeneity. CD, nmr, and modeling analysis indicated that in aqueous media the first extracellular loop peptide E1(107-125), the third intracellular loop peptide I3(231-243), and the carboxyl terminus peptide I4(350-372) were mostly disordered. In contrast, the second extracellular loop peptide E2(191-206) assumed a well-defined structure in aqueous medium and the sixth transmembrane domain peptide receptor M6(252-269, C252A) was highly helical in trifluoroethanol/water (4:1), exhibiting a kink at Pro258.

Electrophoretic behavior of L- and D-alanine-scanning analogs of a yeast tridecapeptide pheromone in a fused-silica capillary.

Electrophoretic behavior of synthetic tridecapeptide diastereomers has been systematically investigated using a series of L-Ala- and D-Ala-scanning analogs of [Nle12] alpha-factor [WHWLQLKPGQP(Nle)Y], a tridecapeptide mating pheromone of Saccharomyces cerevisiae. The effects of buffer pH, buffer concentration, voltage, and temperature on diastereomer separation were tested. Among 13 pairs of diastereomers, those with L-Ala/D-Ala replacement in the middle of the peptide chain exhibited much higher diastereomeric resolution than those with identical replacement near the peptide termini.