Molecular determinants of bioactivity of the Saccharomyces cerevisiae lipopeptide mating pheromone.
The a-factor of Saccharomyces cerevisiae (YIIKGVF-WDPAC(Farnesyl)-OCH3) is a peptide pheromone in which post-translational modification with a farnesyl isoprenoid and carboxyl methyl group is required for export and bioactivity. Truncated and carboxyl-terminal modified analogs of the a-factor were synthesized in order to determine the effect of such modifications on bioactivity. Bioactivity studies on carboxyl-terminal analogs in which the chirality, the cysteine thioether, and the carboxyl ester were varied in an attempt to study the influence of topology on a-factor activity indicate that the hydrophobicity conferred by the farnesyl moiety and not its specific spatial orientation is a key determinant of a-factor potency. Analyses on truncated a-factors suggest that sequential removal of NH2-terminal residues leads to a gradient of potency loss, with some amino acids exhibiting a slightly greater contribution to bioactivity than others. Random oligonucleotide-targeted mutagenesis of the gene encoding a-factor was coupled to a biological screen to identify altered a-factor peptides which are secreted yet exhibit a loss of a-factor bioactivity. Transformants exhibiting this phenotype were examined to identify codon changes presumably responsible for the altered phenotype, thus indicating residues that may contribute significantly to a-factor bioactivity.
Caldwell
G A
GA
Program in Cellular, Molecular, and Developmental Biology, University of Knoxville 37996-0845.
Wang
S H
SH
Xue
C B
CB
Jiang
Y
Y
Lu
H F
HF
Naider
F
F
Becker
J M
JM
D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid; D002855:Chromatography, Thin Layer; D000072235:Mating Factor; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010446:Peptide Fragments; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010641:Phenotype; D011487:Protein Conformation; D012441:Saccharomyces cerevisiae / Q000378:metabolism*
D010446:Peptide Fragments; D010455:Peptides; D000072235:Mating Factor