Legge G B, Kriwacki R W, Chung J, Hommel U, Ramage P, Case D A, Dyson H J, Wright P E
Department of Molecular Biology MB2 and Skaggs Institute for Chemical Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA, 92037, USA.
J Mol Biol. 2000 Feb 4;295(5):1251-64. doi: 10.1006/jmbi.1999.3409.
The interaction between the leukocyte function-associated antigen-1 (LFA-1) and the intercellular adhesion molecule is thought to be mediated primarily via the inserted domain (I-domain) in the alpha-subunit. The activation of LFA-1 is an early step in triggering the adhesion of leukocytes to target cells decorated with intercellular adhesion molecules. There is some disagreement in the literature over the respective roles of conformational changes in the I-domain and of divalent cations (Mg(2+), Mn(2+)) in the activation of LFA-1 for intercellular adhesion molecule binding. X-ray crystallographic structures of the I-domains of LFA-1 and Mac-1 in the presence and absence of cations show structural differences in the C-terminal alpha-helix; this change was proposed to represent the active and inactive conformations of the I-domain. However, more recent X-ray results have called this proposal into question. The solution structure of the Mg(2+) complex of the I-domain of LFA-1 has been determined by NMR methods, using a model-based approach to nuclear Overhauser enhancement spectroscopy peak assignment. The protein adopts the same structure in solution as that of the published I-domain X-ray structures, but the C-terminal region, where the X-ray structures are most different from each other, is different again in the solution structures. The secondary structure of this helix is well formed, but NMR relaxation data indicate that there is considerable flexibility present, probably consisting of breathing or segmental motion of the helix. The conformational diversity seen in the various X-ray structures could be explained as a result of the inherent flexibility of this C-terminal region and as a result of crystal contacts. Our NMR data are consistent with a model where the C-terminal helix has the potential flexibility to take up alternative conformations, for example, in the presence and absence of the intercellular adhesion molecule ligand. The role of divalent cations appears from our results not to be as a direct mediator of a conformational change that alters affinity for the ligand. Rather, the presence of the cation appears to be involved in some other way in ligand binding, perhaps by acting as a bridge to the ligand and by modulation of the charge of the binding surface.
白细胞功能相关抗原-1（LFA-1）与细胞间黏附分子之间的相互作用被认为主要通过α亚基中的插入结构域（I结构域）介导。LFA-1的激活是触发白细胞与装饰有细胞间黏附分子的靶细胞黏附的早期步骤。关于I结构域构象变化和二价阳离子（Mg(2+)、Mn(2+)）在激活LFA-1以结合细胞间黏附分子方面各自的作用，文献中存在一些分歧。LFA-1和Mac-1的I结构域在有和没有阳离子存在时的X射线晶体结构显示C末端α螺旋存在结构差异；这种变化被认为代表了I结构域的活性和非活性构象。然而，最近的X射线结果对这一观点提出了质疑。LFA-1的I结构域的Mg(2+)复合物的溶液结构已通过核磁共振方法确定，采用基于模型的方法进行核Overhauser增强光谱峰归属。该蛋白质在溶液中的结构与已发表的I结构域X射线结构相同，但X射线结构彼此差异最大的C末端区域在溶液结构中又有所不同。该螺旋的二级结构形成良好，但核磁共振弛豫数据表明存在相当大的灵活性，可能由螺旋的呼吸或片段运动组成。各种X射线结构中看到的构象多样性可以解释为该C末端区域固有的灵活性以及晶体接触的结果。我们的核磁共振数据与一个模型一致，即C末端螺旋具有采取替代构象的潜在灵活性，例如，在存在和不存在细胞间黏附分子配体的情况下。从我们的结果来看，二价阳离子的作用似乎不是作为改变对配体亲和力的构象变化的直接介导者。相反，阳离子的存在似乎以某种其他方式参与配体结合，也许是通过作为与配体的桥梁并调节结合表面的电荷。
