Role of the glycoprotein Ib-binding A1 repeat and the RGD sequence in platelet adhesion to human recombinant von Willebrand factor.
To assess the relative importance of the glycoprotein (GP) Ib binding domain and the RGDS binding site in platelet adhesion to isolated von Willebrand factor (vWF) and to collagen preincubated with vWF, we deleted the A1 domain yielding delta A1-vWF and introduced an aspartate-to-glycine substitution in the RGDS sequence by site-directed mutagenesis (RGGS-vWF). Recombinant delta A1-vWF and RGGS-vWF, purified from transfected baby hamster kidney cells, were compared with recombinant wild-type vWF (WT-vWF) in platelet adhesion under static and flow conditions. Purified mutants were coated on glass or on a collagen type III surface and exposed to circulating blood in a perfusion system. Platelet adhesion under static condition, under flow conditions, and in vWF-dependent adhesion to collagen has an absolute requirement for GPIb-vWF interaction. The GPIIb/IIIa-vWF interaction is required for adhesion to coated vWF under flow conditions. Under static condition and vWF-dependent adhesion to collagen, platelet adhesion to RGGS-vWF is similar as to WT-vWF, but platelet spreading and aggregation are abolished.
Lankhof
H
H
Department of Hematology, University Hospital, Utrecht, The Netherlands.
Wu
Y P
YP
Vink
T
T
Schiphorst
M E
ME
Zerwes
H G
HG
de Groot
P G
PG
Sixma
J J
JJ
D000595:Amino Acid Sequence; D001483:Base Sequence; D001792:Blood Platelets / Q000166:cytology*; D015815:Cell Adhesion Molecules / Q000378:metabolism*; D003094:Collagen / Q000378:metabolism; D017931:DNA Primers / Q000737:chemistry; D006801:Humans; D066298:In Vitro Techniques; D008969:Molecular Sequence Data; D009842:Oligopeptides; D010973:Platelet Adhesiveness; D010980:Platelet Membrane Glycoproteins / Q000378:metabolism*; D017354:Point Mutation; D011485:Protein Binding; D011994:Recombinant Proteins; D012212:Rheology; D013329:Structure-Activity Relationship; D014841:von Willebrand Factor / Q000378:metabolism*
D015815:Cell Adhesion Molecules; D017931:DNA Primers; D009842:Oligopeptides; D010980:Platelet Membrane Glycoproteins; D011994:Recombinant Proteins; D014841:von Willebrand Factor; C047981:arginyl-glycyl-aspartic acid; D003094:Collagen