title: Requirement of leucine-rich repeats of glycoprotein (GP) Ibalpha for shear-dependent and static binding of von Willebrand factor to the platelet membrane GP Ib-IX-V complex.
abstract: The platelet glycoprotein (GP) Ib-IX-V complex mediates adhesion to von Willebrand factor (vWf) in (patho)physiologic thrombus formation. The vWf-binding site on GP Ib-IX-V is within the N-terminal 282 residues of GP Ibalpha, which consist of an N-terminal flanking sequence (His-1-Ile-35), 7 leucine-rich repeats (Leu-36-Ala-200), a C-terminal flank (Phe-201-Gly-268), and a sulfated tyrosine sequence (Asp-269-Glu-282). We have used mammalian cell expression of canine-human chimeras of GP Ibalpha, corresponding to precise structural boundaries, to demonstrate the first specific requirement for individual leucine-rich repeats for binding of vWf either induced by a modulator, ristocetin, or under hydrodynamic flow. Implicit in this approach was that the GP Ibalpha chimeras retained a functional conformation, a supposition confirmed by analyzing restoration of function to reversed human-canine chimeras and demonstrating that all chimeras bound vWf activated by botrocetin, a modulator that is indiscriminate between species. Leucine-rich repeats 2, 3, and 4 of GP Ibalpha were identified as being critical for vWf adhesion to GP Ib-IX-V.
authors.affiliation: Hazel and Pip Appel Vascular Biology Laboratory, Baker Medical Research Institute, Melbourne, Australia.
authors.forename: Y
authors.initials: Y
authors.lastname: Shen
authors.forename: G M
authors.initials: GM
authors.lastname: Romo
authors.forename: J F
authors.initials: JF
authors.lastname: Dong
authors.forename: A
authors.initials: A
authors.lastname: Schade
authors.forename: L V
authors.initials: LV
authors.lastname: McIntire
authors.forename: D
authors.initials: D
authors.lastname: Kenny
authors.forename: J C
authors.initials: JC
authors.lastname: Whisstock
authors.forename: M C
authors.initials: MC
authors.lastname: Berndt
authors.forename: J A
authors.initials: JA
authors.lastname: López
authors.forename: R K
authors.initials: RK
authors.lastname: Andrews
mesh_terms: D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D016824:Antigens, Human Platelet / Q000276:immunology; D016466:CHO Cells; D002448:Cell Adhesion; D006224:Cricetinae; D003412:Cricetulus; D003435:Crotalid Venoms / Q000494:pharmacology; D004285:Dogs; D018056:Hemorheology; D006801:Humans; D007930:Leucine / Q000737:chemistry*; D046911:Macromolecular Substances; D010974:Platelet Aggregation / Q000187:drug effects; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D020449:Repetitive Sequences, Amino Acid; D012310:Ristocetin / Q000494:pharmacology; D013314:Stress, Mechanical; D013329:Structure-Activity Relationship; D014841:von Willebrand Factor / Q000378:metabolism*
chemical_list: D000911:Antibodies, Monoclonal; D016824:Antigens, Human Platelet; D003435:Crotalid Venoms; D046911:Macromolecular Substances; D019038:Platelet Glycoprotein GPIb-IX Complex; D011993:Recombinant Fusion Proteins; D014841:von Willebrand Factor; D012310:Ristocetin; C030961:botrocetin; D007930:Leucine