An activated L-selectin mutant with conserved equilibrium binding properties but enhanced ligand recognition under shear flow
An activated L-selectin mutant with conserved equilibrium binding properties but enhanced ligand recognition under shear flow
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DOI：
10.1074/jbc.m001103200
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发表时间：
2000-06-23
影响因子：
4.8
通讯作者：
Alon, R
中科院分区：
文献类型：
--
作者：
Dwir, O;Kansas, GS;Alon, R
选择素介导白细胞在血管壁上的初始束缚和滚动。选择素的粘附是平衡时的配体识别和施加力下的选择素-配体键的机械性质两者的函数。我们描述了一个EGF结构域突变的L-选择素与深刻增强的native L-选择素，但保守的配体特异性。这种突变体，称为LPL，是通过用来自P-选择素的同源结构域取代L-选择素的EGF样结构域而得到的。该突变体结合可溶性碳水化合物L-选择素配体的亲和力与天然L-选择素的亲和力相当，但与所有表面结合的配体的相互作用比天然L-选择素更容易，特别是在高剪切流下。由天然和突变的L-选择素介导的系链在一定范围的剪切应力下表现出相似的寿命，但突变体的成键速率比天然L-选择素对不同的L-选择素配体的成键速率高至少10倍。增强键形成率的突变体与深刻更强的滚动相互作用和减少依赖滚动的阈值剪切应力。这是第一次证明EGF结构域可以调节选择素的凝集素结构域在剪切流下与表面固定化配体的结合，而不影响选择素对可溶性配体的平衡性质。
Selectins mediate the initial tethering and rolling of leukocytes on vessel walls. Adhesion by selectins is a function of both ligand recognition at equilibrium and mechanical properties of the selectin-ligand bond under applied force. We describe an EGF domain mutant of L-selectin with profoundly augmented adhesiveness over that of native L-selectin but conserved ligand specificity. This mutant, termed LPL, was derived by a substitution of the EGF-like domain of L selectin with the homologous domain from P-selectin. The mutant bound soluble carbohydrate L-selectin ligand with affinity comparable with that of native L-selectin but interacted with all surface-bound ligands much more readily than native L-selectin, in particular under elevated shear flow. Tethers mediated by both native and mutant L-selectin exhibited similar lifetimes under a range of shear stresses, but the rate of bond formation by the mutant was at least 10-fold higher than that of native L-selectin toward distinct L-selectin ligands. Enhanced rate of bond formation by the mutant was associated with profoundly stronger rolling interactions and reduced dependence of rolling on a threshold of shear stress. This is the first demonstration that the EGF domain can modulate the binding of the lectin domain of a selectin to surface-immobilized ligands under shear flow without affecting the equilibrium properties of the selectin toward soluble ligands.