Refined crystal structure of the influenza virus N9 neuraminidase-NC41 Fab complex.
Refined crystal structure of the influenza virus N9 neuraminidase-NC41 Fab complex.
复制标题
流感病毒 N9 神经氨酸酶-NC41 Fab 复合物的精制晶体结构。
DOI：
10.1016/0022-2836(92)90687-f
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发表时间：
1992
影响因子：
5.6
通讯作者：
Colman,PM
中科院分区：
文献类型：
--
作者：
Tulip,WR;Varghese,JN;Laver,WG;Webster,RG;Colman,PM
用最小二乘法和模拟退火法对禽流感病毒N9亚型神经氨酸酶与单克隆抗体NC 41的抗原结合片段（Fab）复合物的晶体结构进行了优化，得到了分辨率为8.0 ~ 2.5 μ m的31，846个衍射数据，R因子为0.191。所得模型与理想键长的均方根偏差为0·016 μ m。四分之一的四聚体复合物包括晶体模型，其具有6577个非氢原子，并由神经氨酸酶中的389个蛋白质残基和8个碳水化合物残基组成。Fab轻链中有214个残基，重链中有221个残基。结果表明，在抗原和抗体的相互作用表面上存在着明显的形状互补性，尽管在界面上原子的堆积密度比在蛋白质内部要松一些。类似地，在抗原和抗体之间存在高度的化学互补性，由一个埋藏的盐键、两个溶剂化的盐键和12个氢键介导。而8个片段中的33个神经氨酸酶残基具有899 μ 2的被相互作用（对1·7 μ 1探针）掩埋的表面积，包括两个己糖单元。NC 41 Fab中有17个残基位于5个互补决定区（complementary determining regions，CDR）中，与神经氨酸酶接触，7个片段中的36个抗体残基具有916 μ g/m2的包埋表面积。的接口是更广泛的比三个溶菌酶-Fab复合物的晶体结构已被确定，作为判断掩埋的表面积和接触残基的数量。在复合和未复合的神经氨酸酶结构之间只有很小的差异（小于1·5 μ g），并且在该分辨率和准确度下，这些差异并不明确。五个CDR的主链构象遵循预测的规范结构。轻链和重链的可变结构域之间的界面不像其他Fab中那样广泛，这是由于NC 41中较少的CDR-CDR相互作用。NC 41 Fab轻链上的第一个CDR被定位成使得它可以在空间上阻碍小底物以及大底物接近神经氨酸酶活性位点口袋，NC 41 Fab与从鲸鱼分离的禽样病毒的N9神经氨酸酶之间的复合物（Globicephala malaena）与燕鸥（Anous minutus）N9-NC 41 Fab复合物同晶型结晶，因为14个燕鸥-鲸鱼氨基酸取代不干扰神经氨酸酶-Fab界面或晶体接触。鲸鱼N9-NC 41 Fab晶体结构，来自最终的燕鸥N9复合物，使用分辨率范围为8.0至2.9 μ m的部分数据集进行精修，R因子为0.157，与理想键长的均方根偏差为0.016 μ m。
The crystal structure of the complex between neuraminidase from influenza virus (subtype N9 and isolated from an avian source) and the antigen-binding fragment (Fab) of monoclonal antibody NC41 has been refined by both least-squares and simulated annealing methods to anR-factor of 0·191 using 31,846 diffraction data in the resolution range 8·0 to 2·5 Å. The resulting model has a root-mean-square deviation from ideal bond-length of 0·016 Å. One fourth of the tetrameric complex comprises the crystallographic model, which has 6577 non-hydrogen atoms and consists of 389 protein residues and eight carbohydrate residues in the neuraminidase. 214 residues in the Fab light chain, and 221 residues in the heavy chain. One putative Ca ion buried in the neuraminidase, and 73 water molecules, are also included.A remarkable shape complementarity exists between the interacting surfaces of the antigen and the antibody, although the packing density of atoms at the interface is somewhat looser than in the interior of a protein. Similarly, there is a high degree of chemical complementarity between the antigen and antibody, mediated by one buried salt-link, two solvated salt-links and 12 hydrogen bonds.The antibody-binding site on neuraminidase is discontinuous and comprises five chain segments and 19 residues in contact, whilst 33 neuraminidase residues in eight segments have 899 Å2of surface area buried by the interaction (to a 1·7 Å probe), including two hexose units. Seventeen residues in NC41 Fab lying in five of the six complementarity determining regions (CDRs) make contact with the neuraminidase and 36 antibody residues in seven segments have 916 Å2of buried surface area. The interface is more extensive than those of the three lysozyme-Fab complexes whose crystal structures have been determined, as judged by buried surface area and numbers of contact residues. There are only small differences (less than 1·5 Å) between the complexed and uncomplexed neuraminidase structures and, at this resolution and accuracy, those differences are not unequivocal. The main-chain conformations of five of the CDRs follow the predicted canonical structures. The interface between the variable domains of the light and heavy chains is not as extensive as in other Fabs, due to less CDR-CDR interaction in NC41.The first CDR on the NC41 Fab light chain is positioned so that it could sterically hinder the approach of small as well as large substrates to the neuraminidase active-site pocket, suggesting a possible mechanism for the observed inhibition of enzyme activity by the antibody.The complex between NC41 Fab and N9 neuraminidase from an avial-like virus isolated from a whale (Globicephala malaena) crystallizes isomorphously to the tern (Anous minutus) N9-NC41 Fab complex because the 14 tern-to-whale amino acid substitutions do not perturb either the neuraminidase-Fab interface or crystal contacts. The whale N9-NC41 Fab crystal structure, derived from the final tern N9 complex, was refined using a partial dataset in the resolution range 8·0 to 2·9 Å to anR-factor of 0·157, with a root-mean-square deviation from ideal bond-length of 0·016 Å.