title: Inherited mutations within the calcium-binding sites of the integrin alpha IIb subunit (platelet glycoprotein IIb). Effects of the amino acid side chain and the amino acid position on cation binding.
abstract: To examine the effects of naturally occurring inherited mutations on the ability of the integrin alpha-subunit, also termed glycoprotein IIb (GPIIb), to bind metal ions, we prepared small synthetic peptides that encompassed individual cation-binding domains, and recombinant GPIIb poly peptides that encompassed all four Ca(2+)-binding domains, and examined their interactions with divalent cations by means of Tb3+-luminescence spectroscopy. Replacement of the highly conserved Gly418 residue, located within the flanking region of the fourth Ca(2+)-binding domain of GPIIb, with a negatively charged Asp residue resulted in marked reduction in the ability to bind divalent cations. A variant form of GPIIb with a deletion of two amino acids at the -1 and X positions of the fourth Ca(2+)-binding domain of GPIIb also failed to bind metal ions in a normal manner. In contrast, a Glanzmann mutation at the -1 position of the first Ca(2+)-binding domain of GPIIb had no effect on divalent-cation-binding ability with either synthetic peptides or recombinant GPIIb polypeptides. These data support the hypothesis that the highly conserved Gly normally found 7-8 residues N-terminal to integrin metal-binding domains plays a critical role in the maintenance of the conformation or orientation of surrounding EF-hand structures so that they can effectively interact with and bind divalent cations. Furthermore, inherited mutations at or near the divalent-cation-binding domains of GPIIb do not necessarily exert their biochemical effects by disruption of cation binding, but can interfere with integrin biogenesis in a Ca(2+)-independent manner.
authors.affiliation: Blood Research Institute, Blood Center of Southeastern Wisconsin, Milwauke 53233-2194, USA.
authors.forename: D E
authors.initials: DE
authors.lastname: Jackson
authors.forename: M
authors.initials: M
authors.lastname: Poncz
authors.forename: M T
authors.initials: MT
authors.lastname: Holyst
authors.forename: P J
authors.initials: PJ
authors.lastname: Newman
mesh_terms: D000595:Amino Acid Sequence; D000596:Amino Acids; D001665:Binding Sites; D001667:Binding, Competitive; D002118:Calcium / Q000378:metabolism; D002412:Cations / Q000378:metabolism; D003001:Cloning, Molecular; D004926:Escherichia coli; D014644:Genetic Variation; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D009154:Mutation; D010446:Peptide Fragments / Q000138:chemical synthesis / Q000737:chemistry; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism
chemical_list: D000596:Amino Acids; D002412:Cations; D010446:Peptide Fragments; D019038:Platelet Glycoprotein GPIb-IX Complex; D011994:Recombinant Proteins; D002118:Calcium