Integrin β3 regions controlling binding of murine mAb 7E3:: Implications for the mechanism of integrin αIIbβ3 activation
Integrin β3 regions controlling binding of murine mAb 7E3:: Implications for the mechanism of integrin αIIbβ3 activation
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DOI：
10.1073/pnas.0404201101
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发表时间：
2004-09-07
影响因子：
11.1
通讯作者：
Coller, BS
中科院分区：
文献类型：
--
作者：
Artoni, A;Li, JH;Coller, BS
阿昔单抗是鼠单抗7 E3的衍生物，通过抑制配体与α IIb β 3受体的结合来预防经皮冠状动脉介入治疗的缺血性并发症。在这项研究中，我们确定了控制7 E3结合的整合素β 3区域。在先前研究发现影响7 E3结合的betaA结构域的两个区域中产生鼠/人氨基酸取代：C177-C184环和K125-N133。T182 N取代和K125 Q突变降低了7 E3与人β 3与α IIb复合物的结合。将人C177-C184区和人W129引入鼠β 3是必要的，足以使7 E3与人α IIb/鼠133复合物结合。虽然我们不能排除变构效应，但我们提出7 E3在C177-C184和W129之间结合，它们在晶体结构中彼此在15埃内，并且靠近β 3金属离子依赖性粘附位点。我们以前证明了7 E3比未活化的血小板更快地与活化的血小板结合。因为它已经提出，α IIb β 3的变化，从一个弯曲到一个扩展的构象激活后，我们假设，7 E3结合不太好弯曲比扩展构象。为了支持这一假设，我们发现7 E3与锁定在弯曲构象中的α IIb β 3构建体的结合较差，而解锁构象恢复了7 E3结合。因此，我们的数据是一致的，与alphaIIbbeta 3存在于相互平衡的未活化血小板，和激活导致alphaIIbbeta 3采用更扩展的构象的botted构象。
Abciximab, a derivative of the murine mAb 7E3, protects against ischemic complications of percutaneous coronary interventions by inhibiting ligand binding to the alphaIIbbeta3 receptor. In this study we identified regions on integrin beta3 that control 7E3 binding. Murine/human amino acid substitutions were created in two regions of the betaA domain that previous studies found to influence 7E3 binding: the C177-C184 loop and K125-N133. The T182N substitution and a K125Q mutation reduced 7E3 binding to human beta3 in complex with alphaIIb. The introduction of both the human C177-C184 region and human W129 into murine beta3 was necessary and sufficient to permit 7E3 binding to the human alphaIIb/murine 133 complex. Although we cannot exclude allosteric effects, we propose that 7E3 binds between C177-C184 and W129, which are within 15 Angstrom of each other in the crystal structure and close to the beta3 metal ion-dependent adhesion site. We previously demonstrated that 7E3 binds more rapidly to activated than unactivated platelets. Because it has been proposed that alphaIIbbeta3 changes from a bent to an extended conformation upon activation, we hypothesized that 7E3 binds less well to the bent than the extended conformation. in support of this hypothesis we found that 7E3 bound less well to an alphaIIbbeta3 construct locked in a bent conformation, and unlocking the conformation restored 7E3 binding. Thus, our data are consistent with alphaIIbbeta3 existing in variably bent conformations in equilibrium with each other on unactivated platelets, and activation resulting in alphaIIbbeta3 adopting a more extended conformation.