Fine Structural and Functional Consequences of Deglycosylation of the Platelet Adhesion Receptor GPIb-IX (CD 42b)

Abstract

To investigate the role of the glycosylation of the platelet receptor glycoprotein Ib (GPIb, CD 42b), platelets and purified GPIb were deglycosylated by neuraminidase, O- and N-glycosidases. N-deglycosylation and neuraminic-acid cleavage had little effect on ristocetin and botrocetin-induced platelet agglutination. However, O-deglycosylation reduced the response by approximately 50%, and total deglycosylation (the combination of all three glycosidases) fully abolished the response to ristocetin. Interestingly, binding of von Willebrand Factor (vWF) to purified GPIb in the presence of ristocetin and botrocetin in a standardized microtiter plate assay was not altered by partial or even by total deglycosylation. Electron microscopy indicated that the normally stretched ∼50 nm long molecule was ∼32 nm after N-deglycosylation, ∼20 nm after O-deglycosylation, and reduced in a ∼15 nm long collapse by total deglycosylation. These results suggest that deglycosylation has major structural impacts on GPIb, strongly impairingplatelet-vWF interactions; however, vWF binding toisolated GPIbremains unaffected.