title: Isolation and chemical characterization of two structurally and functionally distinct forms of botrocetin, the platelet coagglutinin isolated from the venom of Bothrops jararaca.
abstract: Two distinct forms of botrocetin, the von Willebrand factor (vWF)-dependent platelet coagglutinin isolated from the venom of the snake Bothrops jararaca, were purified and characterized structurally and functionally. The apparent molecular mass of the one-chain botrocetin was 28 kDa before and 32 kDa after reduction of disulfide bonds, while that of the two-chain botrocetin was 27 kDa before and 15/14.5 kDa after reduction. Amino acid composition of the two species revealed a similar high content of potentially acidic residues (greater than 60 Asx and Glx residues/molecule) but significant differences in the content of Cys and Phe residues. The NH2-terminal sequence of the one-chain botrocetin was Ile-Ile/Val-Ser-Pro-Pro-Val-Cys-Gly-Asn-Glu-. Two constituent polypeptides of the two-chain botrocetin showed similar but different NH2-terminal sequences, distinct from that of the one-chain species: (alpha) Asp-Cys-Pro-Ser-Gly-Trp-Ser-Ser-Tyr-Glu- and (beta) Asp-Cys-Pro-Pro-Asp-Trp-Ser-Ser-Tyr-Glu-. The carbohydrate content of both species was less than 2% of the total mass, and the pI was 4.0-4.1 for the one-chain species, and 4.6, 5.3-5.4, and 7.7-7.8 for the two-chain species. No free sulfhydryl group was detected in each species. Both types of botrocetin were resistant to proteolysis at neutral pH. Incubation of 125I-labeled one-chain botrocetin with the crude venom solution resulted in no detectable structural change. On a weight basis, the two-chain botrocetin was 34 times more active than the one-chain form in promoting vWF binding to platelets.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Department of Blood Transfusion, Nara Medical College, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Fujimura
authors.forename: K
authors.initials: K
authors.lastname: Titani
authors.forename: Y
authors.initials: Y
authors.lastname: Usami
authors.forename: M
authors.initials: M
authors.lastname: Suzuki
authors.forename: R
authors.initials: R
authors.lastname: Oyama
authors.forename: T
authors.initials: T
authors.lastname: Matsui
authors.forename: H
authors.initials: H
authors.lastname: Fukui
authors.forename: M
authors.initials: M
authors.lastname: Sugimoto
authors.forename: Z M
authors.initials: ZM
authors.lastname: Ruggeri
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001792:Blood Platelets / Q000502:physiology; D002241:Carbohydrates / Q000032:analysis; D003435:Crotalid Venoms / Q000302:isolation & purification* / Q000494:pharmacology; D006388:Hemagglutinins / Q000302:isolation & purification*; D006801:Humans; D007700:Kinetics; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D008970:Molecular Weight; D012689:Sequence Homology, Nucleic Acid; D012911:Snakes; D013438:Sulfhydryl Compounds / Q000032:analysis; D014841:von Willebrand Factor / Q000378:metabolism
chemical_list: D000596:Amino Acids; D002241:Carbohydrates; D003435:Crotalid Venoms; D006388:Hemagglutinins; D046911:Macromolecular Substances; D013438:Sulfhydryl Compounds; D014841:von Willebrand Factor; C030961:botrocetin