A two-amino acid insertion in the Cys146- Cys167 loop of the alphaIIb subunit is associated with a variant of Glanzmann thrombasthenia. Critical role of Asp163 in ligand binding.

Honda, Susumu; Tomiyama, Yoshiaki; Shiraga, Masamichi; Tadokoro, Seiji; Takamatsu, Junki; Saito, Hidehiko; Kurata, Yoshiyuki; Matsuzawa, Yūji

doi:10.1172/jci3206

Cited by 48 publications

(58 citation statements)

References 53 publications

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“…In non-I-domain integrin ␣ subunits, particularly the ␣ IIb subunit, several ligand-binding sites have been identified by the characterization of naturally occurring mutations in patients with Glanzmann thrombasthenia as well as mutagenesis analyses. [15][16][17][18][19] However, ligand-binding regions of ␣ v remain elusive. To clarify the critical regions for ligand binding in the ␣ v subunit, we focused on the predicted W3 4-1 loop (C142-C155) and W3 2-3 loop (G172-G181) and investigated their role in ligand binding by alanine-scanning mutagenesis.…”

Section: Discussionmentioning

confidence: 99%

“…Interestingly, many critical residues identified in non-I-domain ␣ subunits have aromatic side chains, suggesting that in addition to oxygenated residues such as D163 in ␣ IIb , 16 aromatic residues are important for ligand binding. 45 The corresponding residues to the ␣ v Y178 in ␣ IIb (Y190), 18 ␣ 4 (Y187), 42,43 and ␣ 5 (F187), 43 and the adjacent residues Y186 and W188 in ␣ 3 40 have been demonstrated to be critical for ligand binding, although the role of W3 2-3 loop in ␣ 3 is still controversial.…”

Section: Discussionmentioning

confidence: 99%

“…Furthermore, the analysis of a Japanese variant of Glanzmann thrombasthenia, KO, and alanine-scanning mutagenesis have shown that D163 in the W3 4-1 loop (cystein [C]146-C167) is essential for ligand binding. 16 In contrast, the single available peptide cross-linking study of ␣ v ␤ 3 showed that 2 distinct linear regions in ␣ v , residues 139-167 and 312-349, cross-link to an arganine-glycine-aspartic acid (RGD) peptide. 21 In this study, we took advantage of the data regarding ligandbinding sites in ␣ IIb and investigated the role in ligand binding of the predicted loops between repeats 2 and 3 (W3 4-1 loop) and within repeat 3 (W3 2-3 loop) in ␣ v .…”

Section: Reprintsmentioning

confidence: 99%

“…14 However, recent characterization of molecular defects in Glanzmann thrombasthenia and mutagenesis studies demonstrated several discontinuous residues important for ligand binding: proline (P)145, D163, L183, G184, tyrosine (Y)189, Y190, phenylalanine (F)191, G193, and D224. [15][16][17][18][19] Springer has proposed that the 7 N-terminal sequence repeats of integrin ␣ subunits are folded into a ␤-propeller domain. 20 The proposed domains contain seven 4-stranded ␤-sheets (W1-W7) arranged in a torus around a pseudosymmetry axis.…”

Section: Introductionmentioning

confidence: 99%

“…To introduce single alanine substitutions into ␣ v , overlap extension polymerase chain reaction (PCR) was carried out as previously described. 16 For example, to generate the Y1783A (Y178A) ␣ v mutant, we synthesized mismatched sense primer ␣ v 178A-s, 5Ј-GGTCCTGGTAGCTTTGCATG-GCAAGGTCAGC-3Ј (sense, nucleotides [nt] 648-678; mismatched sequences underlined) and antisense primer ␣ v 178A-as, 5Ј-GCTGACCTTGC-CATGCAAAGCTACCAGGACC-3Ј (antisense, nt 678-648; mismatched sequences underlined), which were constructed based on the published sequence. 31 PCR was performed by using ␣ v cDNA as a template and primers ␣ v 258-s, 5Ј-GCAAACACCACCCAGCC-3Ј (sense, nt 258-274) and ␣ v 178A-as, or primers ␣ v 178A-s and ␣ v 952-as, 5Ј-GCAGCCATCT-GCTCGCCAG-3Ј (antisense, nt 952-934).…”

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Ligand binding to integrin αvβ3requires tyrosine 178 in the αv subunit

Honda

Tomiyama

Pampori

et al. 2001

Blood

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Integrin ␣ v ␤ 3 has been implicated in angiogenesis and other biological processes. However, the ligand-binding sites in ␣ v , a non-I-domain ␣ subunit, remain to be identified. Recently in ␣ IIb , the other partner of the ␤ 3 subunit, several discontinuous residues important for ligand binding were identified in the predicted loops between repeats 2 and 3 (W3 4-1 loop) and within repeat 3 (W3 2-3 loop). Based on these findings, alaninescanning mutagenesis in 293 cells was used to investigate the role of these loops (cys- teine IntroductionIntegrins are a large family of ␣␤ heterodimeric adhesion receptors that is often subdivided into groups based on 8 known integrin ␤ subunits. 1 The ␤ 3 -integrin subfamily is composed of ␣ v ␤ 3 , originally identified as the vitronectin receptor, and ␣ IIb ␤ 3 , a plateletspecific receptor for fibrinogen and von Willebrand factor. ␣ IIb ␤ 3 plays a crucial role in platelet aggregation, normal hemostasis, and pathological thrombus formation. 2 On the other hand, ␣ v ␤ 3 is expressed in a number of tissues, including platelets, endothelial cells, vascular smooth muscle cells, and osteoclasts, and it plays a key role in angiogenesis and bone resorption. 3,4 The ␣ IIb and ␣ v subunits are homologous and 36% identical in primary amino acid sequence, 5 and most ligands that bind to ␣ IIb ␤ 3 , including fibrinogen, von Willebrand factor, and vitronectin, also bind to ␣ v ␤ 3 . However, there are some distinctive features between these 2 integrins. 3 First, the ␣ IIb subunit has been found only in combination with ␤ 3 , whereas ␣ v can associate with at least 5 ␤ subunits (␤ 1 , ␤ 3 , ␤ 5 , ␤ 6 , and ␤ 8 ). 6 Second, some ligands, such as osteopontin, matrix metalloproteinase-2, and adenovirus penton base, bind to ␣ v ␤ 3 but not to ␣ IIb ␤ 3 . Third, treatment of ␣ IIb ␤ 3 with ethylenediamine tetraacetic acid (EDTA) at 37°C dissociates the complex into its individual subunits; ␣ v ␤ 3 remains a heterodimer. Finally, the ligand-binding function of ␣ v ␤ 3 , but not ␣ IIb ␤ 3 , is suppressed by calcium (Ca 2ϩ ). 7 Generally, all integrins require divalent cations for ligand recognition, and multiple residues important for ligand binding have been identified on both ␣ and ␤ subunits. 8 The N-terminal region of integrin ␣ subunits has 7 repeats of homologous sequences of about 60 amino acid residues. Some integrin ␣ subunits (eg, ␣ 2 , ␣ L , and ␣ M ) contain an inserted domain of about 200 amino acids residues (the I-domain) between the second and the third repeats in the ␣ subunit, which is critically involved in ligand binding. 9,10 The crystal structure of the I-domain has been determined, and the metal ion-dependent adhesion site (MIDAS) motif that contributes to cation binding, as well as ligand binding, has been clarified. 11 Interestingly, a MIDAS-like motif essential for the ligand-binding function was also identified in integrin ␤ subunits. 12,13 On the other hand, integrin ␣ subunits, such as ␣ v , ␣ IIb , and ␣ 4 , do not have the I-domain. The structural basis for the i...

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Reprintsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

mentioning

confidence: 99%

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Ligand binding to integrin αvβ3requires tyrosine 178 in the αv subunit

Honda

Tomiyama

Pampori

et al. 2001

Blood

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Platelet Disorders

and¹,

McCrae²

2005

Molecular Hematology

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Molecular dynamics simulations corroborate recombinant expression studies carried out on three αIIb β‐propeller mutations reported in Indian Glanzmann thrombasthenia patients

Chandrasekaran,

Vishal,

Arora

et al. 2023

J of Cellular Biochemistry

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Mutations in the αIIb β‐propeller domain have long been known to disrupt heterodimerization and intracellular trafficking of αIIbβ3 complexes leading to diminished surface expression and/or function, resulting in Glanzmann thrombasthenia. Our previous study on three β‐propeller mutations, namely G128S, S287L, and G357S, showed variable defects in protein transport correlated with the patient's clinical phenotypes. Pulse‐chase experiments revealed differences in αIIbβ3 complex maturation among the three mutations. Hence, the current study aims to correlate conformational changes caused by each one of them. Evolutionary conservation analysis, stability analysis, and molecular dynamics simulations of the three mutant structures were carried out. Stability analysis revealed that, while G128S and G357S mutations destabilized the β‐propeller structure, S287L retained the stability. Wild‐type and mutant β‐propeller structures, when subjected to molecular dynamics simulations, confirmed that G128S and G357S were both destabilizing in nature when compared with the wild‐type and S287L based on several parameters studied, like RMSD, RMSF, Rg, FEL, PCA, secondary structure, and hydrogen bonds. In our previous study, we demonstrated that mutant S287L αIIbβ3 complexes were more stable than the wild‐type αIIbβ3 complexes, as evidenced in pulse‐chase experiments. These findings corroborate variable intracellular fates of mutant αIIbβ3 complexes as a result of these β‐propeller mutations.

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A two-amino acid insertion in the Cys146- Cys167 loop of the alphaIIb subunit is associated with a variant of Glanzmann thrombasthenia. Critical role of Asp163 in ligand binding.

Cited by 48 publications

References 53 publications

Ligand binding to integrin αvβ3requires tyrosine 178 in the αv subunit

Ligand binding to integrin αvβ3requires tyrosine 178 in the αv subunit

Platelet Disorders

Molecular dynamics simulations corroborate recombinant expression studies carried out on three αIIb β‐propeller mutations reported in Indian Glanzmann thrombasthenia patients

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