title: A naturally occurring Tyr143His alpha IIb mutation abolishes alpha IIb beta 3 function for soluble ligands but retains its ability for mediating cell adhesion and clot retraction: comparison with other mutations causing ligand-binding defects.
abstract: The molecular basis for the interaction between a prototypic non-I-domain integrin, alpha(IIb)beta(3), and its ligands remains to be determined. In this study, we have characterized a novel missense mutation (Tyr143His) in alpha(IIb) associated with a variant of Glanzmann thrombasthenia. Osaka-12 platelets expressed a substantial amount of alpha(IIb)beta(3) (36%-41% of control) but failed to bind soluble ligands, including a high-affinity alpha(IIb)beta(3)-specific peptidomimetic antagonist. Sequence analysis revealed that Osaka-12 is a compound heterozygote for a single (521)T>C substitution leading to a Tyr143His substitution in alpha(IIb) and for the null expression of alpha(IIb) mRNA from the maternal allele. Given that Tyr143 is located in the W3 4-1 loop of the beta-propeller domain of alpha(IIb), we examined the effects of Tyr143His or Tyr143Ala substitution on the expression and function of alpha(IIb)beta(3) and compared them with KO (Arg-Thr insertion between 160 and 161 residues of alpha(IIb)) and with the Asp163Ala mutation located in the same loop by using 293 cells. Each of them abolished the binding function of alpha(IIb)beta(3) for soluble ligands without disturbing alpha(IIb)beta(3) expression. Because immobilized fibrinogen and fibrin are higher affinity/avidity ligands for alpha(IIb)beta(3), we performed cell adhesion and clot retraction assays. In sharp contrast to KO mutation and Asp163Ala alpha(IIb)beta(3), Tyr143His alpha(IIb)beta(3)-expressing cells still had some ability for cell adhesion and clot retraction. Thus, the functional defect induced by Tyr143His alpha(IIb) is likely caused by its allosteric effect rather than by a defect in the ligand-binding site itself. These detailed structure-function analyses provide better understanding of the ligand-binding sites in integrins.
authors.affiliation: Department of Internal Medicine and Molecular Science, Graduate School of Medicine, Suita, Osaka University, Japan.
authors.forename: Teruo
authors.initials: T
authors.lastname: Kiyoi
authors.forename: Yoshiaki
authors.initials: Y
authors.lastname: Tomiyama
authors.forename: Shigenori
authors.initials: S
authors.lastname: Honda
authors.forename: Seiji
authors.initials: S
authors.lastname: Tadokoro
authors.forename: Morio
authors.initials: M
authors.lastname: Arai
authors.forename: Hirokazu
authors.initials: H
authors.lastname: Kashiwagi
authors.forename: Satoru
authors.initials: S
authors.lastname: Kosugi
authors.forename: Hisashi
authors.initials: H
authors.lastname: Kato
authors.forename: Yoshiyuki
authors.initials: Y
authors.lastname: Kurata
authors.forename: Yuji
authors.initials: Y
authors.lastname: Matsuzawa
mesh_terms: D000328:Adult; D000494:Allosteric Regulation; D019943:Amino Acid Substitution; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism; D003018:Clot Retraction; D003062:Codon / Q000235:genetics; D004151:Dipeptides / Q000494:pharmacology; D005260:Female; D005340:Fibrinogen / Q000378:metabolism; D006579:Heterozygote; D006801:Humans; D008024:Ligands; D020125:Mutation, Missense; D010973:Platelet Adhesiveness / Q000502:physiology*; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000737:chemistry / Q000502:physiology*; D040201:Platelet Membrane Glycoprotein IIb / Q000737:chemistry / Q000235:genetics*; D017354:Point Mutation; D012150:Polymorphism, Restriction Fragment Length; D011485:Protein Binding; D017434:Protein Structure, Tertiary / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis; D011993:Recombinant Fusion Proteins / Q000502:physiology; D012995:Solubility; D013329:Structure-Activity Relationship; D013915:Thrombasthenia / Q000097:blood / Q000235:genetics*; D014162:Transfection
chemical_list: D003062:Codon; D004151:Dipeptides; D008024:Ligands; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D040201:Platelet Membrane Glycoprotein IIb; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; C103087:((4-(4-amidinophenoxy)butanoyl)aspartyl)valine; D005340:Fibrinogen