title: Glycoprotein Ibalpha forms disulfide bonds with 2 glycoprotein Ibbeta subunits in the resting platelet.
abstract: It is widely accepted that glycoprotein (GP) Ib contains one Ibalpha and one Ibbeta subunit that are connected by a disulfide bond. It is unclear which Cys residue in Ibalpha, C484 or C485, forms the disulfide bond with Ibbeta. Using mutagenesis studies in transfected Chinese hamster ovary (CHO) cells, we found that both C484 and C485 formed a disulfide bond with C122 in Ibbeta. In the context of isolated peptides containing the Ibalpha or Ibbeta transmembrane domain and nearby Cys residue, C484 and C485 in the Ibalpha peptide were both capable of forming a disulfide bond with the Ibbeta peptide. Furthermore, coimmunoprecipitation of epitope-tagged subunits showed that at least 2 Ibbeta subunits but only 1 Ibalpha and 1 IX subunit were present in the GP Ib-IX complex. Finally, the size difference between GP Ib from transfected CHO cells and human platelets was attributed to a combination of sequence polymorphism and glycosylation difference in Ibalpha, not the number of Ibbeta subunits therein. Overall, these results demonstrate that Ibalpha is covalently connected to 2 Ibbeta subunits in the resting platelet, necessitating revision of the subunit stoichiometry of the GP Ib-IX-V complex. The alphabeta2 composition in GP Ib may provide the basis for possible disulfide rearrangement in the receptor complex.
authors.affiliation: Center for Membrane Biology, Department of Biochemistry and Molecular Biology, The University of Texas Health Science Center at Houston, MSB 6.130, 6431 Fannin St, Houston, TX 77030, USA.
authors.forename: Shi-Zhong
authors.initials: SZ
authors.lastname: Luo
authors.forename: Xi
authors.initials: X
authors.lastname: Mo
authors.forename: Vahid
authors.initials: V
authors.lastname: Afshar-Kharghan
authors.forename: Sankaranarayanan
authors.initials: S
authors.lastname: Srinivasan
authors.forename: José A
authors.initials: JA
authors.lastname: López
authors.forename: Renhao
authors.initials: R
authors.lastname: Li
mesh_terms: D000818:Animals; D001792:Blood Platelets / Q000737:chemistry / Q000378:metabolism*; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D003545:Cysteine / Q000737:chemistry / Q000235:genetics; D004220:Disulfides / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D016297:Mutagenesis, Site-Directed; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014162:Transfection
chemical_list: D004220:Disulfides; D019038:Platelet Glycoprotein GPIb-IX Complex; D003545:Cysteine