Weis W I, Brünger A T, Skehel J J, Wiley D C
Howard Hughes Medical Institute, Yale University, New Haven, CT 06511.
J Mol Biol. 1990 Apr 20;212(4):737-61. doi: 10.1016/0022-2836(90)90234-D.
We have applied the method of simulated annealing to the refinement of the 3 A resolution crystal structure of the influenza virus hemagglutinin glycoprotein, using the program X-PLOR. Two different methods were introduced into X-PLOR to treat the non-crystallographic symmetry present in this and in other crystal structures. In the first, only the unique protomer atoms are refined; by application of the non-crystallographic symmetry operators to the protomer atoms, the X-ray structure factor derivatives are effectively averaged, and a non-bonded energy term models the interactions of the protomer with its neighbors in the oligomer without explicit refinement of the other protomers in the crystallographic asymmetric unit. In the second method, the entire asymmetric unit is refined, but an effective energy term is added to the empirical energy that restrains symmetry-related atomic positions to their average values after least-squares superposition. Several other modifications and additions were made to previously published X-PLOR protocols, including weighting of the X-ray terms, maintenance of the temperature of the molecular dynamics simulation, treatment of charged groups, changes in the values of certain empirical energy parameters, and the use of N-linked carbohydrate empirical energy parameters. The hemagglutinin refinement proceeded in several stages. An initial round of simulated annealing of the monomer was followed by rigid-body refinement of the 3-fold non-crystallographic symmetry axis position and a second round of monomer refinement. A third round was performed on the trimer using non-crystallographic symmetry restraints in all regions except those in lattice contacts showing obvious derivations from 3-fold symmetry. The refinement was completed with several rounds of conventional positional and isotropic temperature factor refinement needed to correct bad model geometry introduced by high-temperature molecular dynamics in regions of weak electron density. This structure was then used as the basis for refinement of three crystallographically isomorphous hemagglutinin structures, including complexes with the influenza virus receptor, sialic acid. Model geometry comparable to well-refined high-resolution structures was obtained with relatively little manual intervention, demonstrating the ability of simulated annealing refinement to produce highly idealized structures at moderate resolution.
我们使用X-PLOR程序，将模拟退火方法应用于流感病毒血凝素糖蛋白3埃分辨率晶体结构的精修。两种不同的方法被引入X-PLOR中，以处理该晶体结构及其他晶体结构中存在的非晶体学对称性。第一种方法是，仅对唯一的原体原子进行精修；通过将非晶体学对称算符应用于原体原子，X射线结构因子导数得到有效平均，并且一个非键能项对原体与其在寡聚体中相邻原子的相互作用进行建模，而无需对晶体学不对称单元中的其他原体进行显式精修。在第二种方法中，对整个不对称单元进行精修，但在经验能量中添加一个有效能项，该能项在最小二乘叠加后将与对称相关的原子位置限制在其平均值。对先前发表的X-PLOR协议进行了其他一些修改和补充，包括对X射线项进行加权、维持分子动力学模拟的温度、处理带电基团、改变某些经验能量参数的值以及使用N-连接碳水化合物经验能量参数。血凝素的精修分几个阶段进行。首先对单体进行一轮模拟退火，随后对3重非晶体学对称轴位置进行刚体精修，然后进行第二轮单体精修。第三轮是对三聚体进行的，在除了晶格接触中显示出明显偏离3重对称性的区域之外的所有区域使用非晶体学对称约束。通过几轮常规的位置和各向同性温度因子精修完成精修，以校正弱电子密度区域中高温分子动力学引入的不良模型几何结构。然后将该结构用作精修三种晶体学同构血凝素结构的基础，包括与流感病毒受体唾液酸的复合物。在相对较少的人工干预下获得了与精修良好的高分辨率结构相当的模型几何结构，这证明了模拟退火精修在中等分辨率下产生高度理想化结构的能力。
