Expression and secretion of biologically active echistatin in Saccharomyces cerevisiae.
A synthetic gene coding for a platelet aggregation inhibitor, echistatin (ECS), was inserted into a Saccharomyces cerevisiae expression vector utilizing the alpha-mating factor pre-pro leader sequence and galactose-inducible promoter, GAL10. Cleavage of the pre-pro leader sequence in vivo results in the secretion of a properly processed recombinant ECS with the native N-terminal glutamic acid residue. Recombinant ECS was recovered from yeast supernatants and purified by reverse phase high performance liquid chromatography. Recombinant ECS expressed and purified from yeast was identical to native ECS in its ability to inhibit platelet aggregation.
Jacobson
M A
MA
Department of Pharmacology, Merck Sharp and Dohme Research Laboratories, West Point, PA 19486.
Forma
F M
FM
Buenaga
R F
RF
Hofmann
K J
KJ
Schultz
L D
LD
Gould
R J
RJ
Friedman
P A
PA
D000595:Amino Acid Sequence; D001483:Base Sequence; D010450:Endopeptidases / Q000235:genetics; D005813:Genes, Synthetic; D005822:Genetic Vectors; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins; D000072235:Mating Factor; D008565:Membrane Proteins; D008969:Molecular Sequence Data; D010455:Peptides / Q000235:genetics; D010675:Pheromones / Q000096:biosynthesis; D010974:Platelet Aggregation; D010975:Platelet Aggregation Inhibitors / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000494:pharmacology; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000235:genetics*; D012697:Serine Endopeptidases; D014757:Viper Venoms / Q000096:biosynthesis / Q000235:genetics* / Q000494:pharmacology
D036341:Intercellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D010455:Peptides; D010675:Pheromones; D010975:Platelet Aggregation Inhibitors; D011994:Recombinant Proteins; D014757:Viper Venoms; C057986:echistatin; D000072235:Mating Factor; D010450:Endopeptidases; D012697:Serine Endopeptidases; C026515:type I signal peptidase