title: "Insularin, a disintegrin from Bothrops insularis venom: inhibition of platelet aggregation and endothelial cell adhesion by the native and recombinant GST-insularin proteins".
abstract: Insularin (INS) was obtained from Bothrops insularis venom by reversed-phase high-performance liquid chromatography using a C(18) column and characterized as a disintegrin by peptide mass fingerprint and inhibition of ADP-induced platelet aggregation. A cDNA coding for P-II a metalloproteinase/disintegrin was cloned from a cDNA library from B. insularis venom glands. The deduced protein sequence possesses 73 amino acid residues, including the N-terminal, internal peptides of native insularin, the ARGDNP-sequence and 12 cysteines in a conserved alignment. This cDNA fragment was subcloned in the pGEX-4T-1 vector and expressed in a prokaryotic expression system as a fusion protein with glutathione S-transferase (GST-INS). Both native and recombinant insularin inhibited ADP-induced platelet aggregation and endothelial cells (HUVEC) adhesion with similar activities indicating that GST-INS folded correctly and preserved the integrin-binding loop. Insularin may be a tool in studies that involve platelets and endothelial cell adhesion dependent on alphaIIbeta3 and alphavbeta3 integrins.
authors.affiliation: Laboratório de Imunopatologia, Instituto Butantan, Av Vital Brazil 1500, 05503-000, São Paulo, SP, Brazil. dellacasa@butantan.gov.br
authors.forename: Maisa Splendore
authors.initials: MS
authors.lastname: Della-Casa
authors.forename: Inácio
authors.initials: I
authors.lastname: Junqueira-de-Azevedo
authors.forename: Diego
authors.initials: D
authors.lastname: Butera
authors.forename: Patrícia Bianca
authors.initials: PB
authors.lastname: Clissa
authors.forename: Daiana S
authors.initials: DS
authors.lastname: Lopes
authors.forename: Solange M T
authors.initials: SM
authors.lastname: Serrano
authors.forename: Daniel C
authors.initials: DC
authors.lastname: Pimenta
authors.forename: Geraldo S
authors.initials: GS
authors.lastname: Magalhães
authors.forename: Paulo Lee
authors.initials: PL
authors.lastname: Ho
authors.forename: Ana Maria
authors.initials: AM
authors.lastname: Moura-da-Silva
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D017837:Bothrops; D002448:Cell Adhesion / Q000187:drug effects; D002478:Cells, Cultured; D003001:Cloning, Molecular; D003435:Crotalid Venoms / Q000096:biosynthesis / Q000737:chemistry* / Q000494:pharmacology; D019483:Disintegrins / Q000096:biosynthesis / Q000737:chemistry / Q000494:pharmacology*; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects*; D005982:Glutathione Transferase / Q000378:metabolism*; D006801:Humans; D007231:Infant, Newborn; D008969:Molecular Sequence Data; D010449:Peptide Mapping; D010974:Platelet Aggregation / Q000187:drug effects*; D010975:Platelet Aggregation Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000494:pharmacology*; D014471:Umbilical Veins / Q000166:cytology
chemical_list: D003435:Crotalid Venoms; D019483:Disintegrins; D010975:Platelet Aggregation Inhibitors; D011993:Recombinant Fusion Proteins; C557217:insularin, Bothrops insularis; D005982:Glutathione Transferase