title: Inhibition of platelet aggregation by grafting RGD and KGD sequences on the structural scaffold of small disulfide-rich proteins.
abstract: Disintegrins represent a group of disulfide-rich peptides ranging in size from 41 to over 80 residues and are antagonists of several integrin receptors. Disintegrins containing an RGD or KGD sequence are potent inhibitors of platelet aggregation as they block the binding of fibrinogen to alpha(IIb)beta(3) integrin. The high affinity binding to alpha(IIb)beta(3) in comparison to short linear peptides has been attributed to the localisation of the RGD or KGD sequence within a defined three-dimensional structure. Cystine knot microproteins are members of another family of small disulfide-rich peptides that consist of only 28-40 amino acid residues. They display numerous biological activities depending on the peptide sequence of loop regions that are fixed on a structural scaffold that is stabilised by three knot-forming disulfide bonds. In the present study we grafted RGD and KGD containing peptide sequences with seven and 11 amino acids, respectively, into two cystine knot microproteins, the trypsin inhibitor EETI-II and the melanocortin receptor binding domain of the human agouti-related protein AGRP, as well as into the small disintegrin obtustatin. The engineered proteins were much more potent to inhibit the fibrinogen binding, alpha(IIb)beta(3) activation and platelet aggregation when compared to the grafted peptides. Differences that were observed between the engineered proteins indicate the importance of the structural scaffold and the amino acids neighbouring the grafted peptide sequences.
authors.affiliation: Clinic for Anaesthesiology and Intensive Care Medicine, University Hospital, Jena, Germany.
authors.forename: Sandy
authors.initials: S
authors.lastname: Reiss
authors.forename: Matthias
authors.initials: M
authors.lastname: Sieber
authors.forename: Volker
authors.initials: V
authors.lastname: Oberle
authors.forename: Alexander
authors.initials: A
authors.lastname: Wentzel
authors.forename: Peter
authors.initials: P
authors.lastname: Spangenberg
authors.forename: Ralf
authors.initials: R
authors.lastname: Claus
authors.forename: Harald
authors.initials: H
authors.lastname: Kolmar
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Lösche
mesh_terms: D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D047168:Cystine Knot Motifs / Q000502:physiology; D019483:Disintegrins / Q000737:chemistry / Q000494:pharmacology*; D005340:Fibrinogen / Q000737:chemistry; D005434:Flow Cytometry; D006801:Humans; D009842:Oligopeptides / Q000235:genetics / Q000494:pharmacology; D010940:Plant Proteins / Q000235:genetics; D010974:Platelet Aggregation / Q000187:drug effects* / Q000502:physiology; D010975:Platelet Aggregation Inhibitors / Q000737:chemistry / Q000494:pharmacology*; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000502:physiology*; D039341:Receptors, Fibrinogen / Q000502:physiology; D014757:Viper Venoms / Q000737:chemistry
chemical_list: D019483:Disintegrins; D009842:Oligopeptides; D010940:Plant Proteins; D010975:Platelet Aggregation Inhibitors; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D039341:Receptors, Fibrinogen; D014757:Viper Venoms; C474313:obtustatin, Vipera lebetina obtusa; C063950:trypsin inhibitor EETI II protein, plant; C047981:arginyl-glycyl-aspartic acid; D005340:Fibrinogen