Study of nucleic acid structure by novel NMR methods

通过新型 NMR 方法研究核酸结构

• 批准号: 7734035
• 负责人: Ad - Bax
• 金额: $ 30.49万
• 依托单位: NATIONAL INSTITUTE OF DIABETES AND DIGESTIVE AND KIDNEY DISEASES
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7734035
• 关键词: Agreement; Algorithms; Bacteriophage Pf1; Base Pairing; Behavior; Class; Compatible; Complement; Computer software; Coupling; Crystallization; Data; Ensure; Exhibits; Homology Modeling; Measures; Messenger RNA; Methods; Modeling; Movement; Nucleic Acids; Phenylalanine-Specific tRNA; Procedures; Proteins; Relative (related person); Residual state; Roentgen Rays; Sampling; Solutions; Speed; Structure; Transfer RNA; Upper arm; Validation; Valine-Specific tRNA; Vertebral column; Yeasts; base; data structure; magnetic field; molecular shape; novel; nucleic acid structure; restraint; ribose phosphate; statistics

A procedure has been developed for refinement of homology models by addition of sparse experimental data. The method is demonstrated for determining the structure of E.Coli tRNAVal, originally modeled after the X-ray structure of yeast tRNAPhe, but refined using experimental residual dipolar coupling (RDC) and small angle X-ray scattering (SAXS) data. A spherical sampling algorithm has been developed for refinement against SAXS data that does not require a globbic approximation, which is particularly important for nucleic acids where such approximations are less appropriate. Substantially higher speed of the algorithm also makes its application favorable for proteins. In addition to the SAXS data, the structure refinement employed a sparse set of NMR data consisting of 24 imino N-HN RDCs measured with Pf1 phage alignment, and 20 imino N-HN RDCs obtained from magnetic field dependent alignment of tRNAVal. The refinement strategy aims to largely retain the local geometry of the 58% identical tRNAPhe by ensuring that the atomic coordinates for short, overlapping segments of the ribose-phosphate backbone and the conserved base pairs remain close to those of the starting model. Local coordinate restraints are enforced using the non-crystallographic symmetry (NCS) term in the XPLOR-NIH or CNS software package, while still permitting modest movements of adjacent segments. The RDCs mainly drive the relative orientation of the helical arms, whereas the SAXS restraints ensure an overall molecular shape compatible with experimental scattering data. The resulting structure exhibits good cross-validation statistics (jack-knifed Qfree = 14% for the Pf1 RDCs, compared to 25% for the starting model) and exhibits a larger angle between the two helical arms than observed in the X-ray structure of tRNAPhe, in agreement with previous NMR-based tRNAVal models.

通过添加稀疏的实验数据，已经开发了一种程序来完善同源模型。证明了该方法用于确定最初以酵母trnaphe的X射线结构建模的大肠杆菌的结构，但使用实验性残留偶性偶联（RDC）和小角度X射线散射（SAXS）数据进行了精制。已经开发出一种球形采样算法，以对不需要球近似的SAXS数据进行完善，这对于此近似值不太合适的核酸尤其重要。该算法的速度基本上也使其适用于蛋白质。 除SAXS数据外，该结构的完善还采用了一组稀疏的NMR数据，该数据由24个Imino N-HN RDC组成，这些NMR数据由PF1噬菌体排列测量，并从依赖于Trnaval的磁场的磁场获得的20个Imino N-HN RDC。改进策略旨在通过确保对核糖磷酸核磷酸核糖链链的短片段和保守的碱基对的原子坐标和保守的碱基对保持接近起始模型的原子坐标，从而在很大程度上保留了58％相同的Trnaphe的局部几何形状。使用Xplor-NIH或CNS软件包中的非结晶对称性（NCS）项来强制局部坐标约束，同时仍允许相邻段的适度运动。 RDCS主要驱动螺旋臂的相对取向，而SAXS约束确保了与实验散射数据兼容的总体分子形状。所得的结构表现出良好的交叉验证统计（PF1 RDC的JackKnife QFREE = 14％，而起始模型为25％），并且在TRNAPHE的X射线结构中表现出的两个螺旋臂之间的角度比以前的NMR基于NMR的TRNAVAL模型一致。