Characterization of novel peptide agonists of the alpha mating factor of Saccharomyces cerevisiae.
Alpha-factor [WHWLQLKPGQPMY], a secreted tridecapeptide pheromone, is required for mating between the a- and alpha-haploid mating types of Saccharomyces cerevisiae (MATa, MATalpha). New analogues of alpha-factor were synthesized and evaluated by morphogenesis assays and receptor binding studies. The Y(0)Nle(12)F(13) analogue [YWHWLQLKPGQPNleF] (MFN5) caused growth arrest and morphological alteration in MATa cells in a fashion identical to that of the native pheromone. Binding of (125)I-labeled MFN5 was saturable, and reversible as shown by equipotent label displacement by MFN5 and native alpha-mating factor. Scatchard analysis of equilibrium binding data on plasma membranes and intact cells indicated the existence of a single high-affinity binding site (K(d) = 6.4 x 10(-8)). Specific binding of (125)I-labeled MFN5 was significantly reduced by guanosine nucleotides. Affinity cross-linking of (125)I-labeled MFN5 to MATa cell membranes identified a specifically labeled 49-kDa protein. The novel synthetic alpha-factor analogue MFN5 can be easily iodinated and used as a probe for the alpha-factor receptor.
Siegel
E G
EG
Laboratory of Molecular Gastroenterology, Gastrointestinal Unit, 1st Department of Medicine, Christian-Albrechts-University of Kiel, Kiel, 24105, Germany.
Günther
R
R
Schäfer
H
H
Fölsch
U R
UR
Schmidt
W E
WE
D000345:Affinity Labels; D000595:Amino Acid Sequence; D001665:Binding Sites; D001681:Biological Assay / Q000379:methods; D002462:Cell Membrane / Q000378:metabolism; D003432:Cross-Linking Reagents; D007457:Iodine Radioisotopes; D000072235:Mating Factor; D008969:Molecular Sequence Data; D010455:Peptides / Q000819:agonists* / Q000032:analysis* / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology; D012441:Saccharomyces cerevisiae / Q000737:chemistry* / Q000187:drug effects; D017386:Sequence Homology, Amino Acid
D000345:Affinity Labels; D003432:Cross-Linking Reagents; D007457:Iodine Radioisotopes; D010455:Peptides; D000072235:Mating Factor