NEW METHODS OF BIOMOLECULAR STRUCTURE DETERMINATION

x射线衍射或晶体学以其独特的能力揭示了广泛的生物医学上重要分子的原子或近原子结构，是现代结构生物学的基石。作为最近基因组研究的最新进展，现在需要对数百种新蛋白质进行有效的x射线晶体学分析。该项目解决了晶体学的关键步骤之一，即所谓的“相位问题”的解决方案。该项目之前的工作已经导致了一种强大的直接方法算法的发展，称为Shake-and-Bake，以及一个计算机程序SnB，可以解决多达1300个独特的非氢原子结构的相位问题。含有多达80个蛋氨酸残基的更大的蛋白质的硒亚结构也可以求解，只要这些残基被硒代蛋氨酸取代。然后，亚结构可以用作引导来定位蛋白质分子的其余部分。现在的总体目标是扩展Shake-and- Bake算法的功能和范围。SnB解决非常大的硒代蛋氨酸亚结构（bbb100 Se位点）的能力将通过添加掩蔽函数来增强，以允许使用有关分子包膜的先验信息或由非晶体对称提供的限制。将制定利用参考光束衍射信息的策略，以促进成功的低分辨率应用。将开发实用的直接方法算法，以最大限度地利用单导数衍射数据（SIR）或具有异常散射的单波长衍射数据（SAS）。phase程序包中的概念和例程将用于整合蛋白质晶体学中常用的其他信息和技术的直接方法，并为可解释的蛋白质电子密度图的自动化生产创造途径。最后，瑞士央行网站http://www.hwi.buffalo.edu/SnB/将继续用于方案传播和用户教育。

X-ray diffraction or crystallography, with its unique ability to reveal the atomic or near-atomic structures of a wide range of biomedically important molecules, is the cornerstone of modern structural biology. As a resent of recent genome studies, there is now a need for efficient X-ray crystallographic analysis of hundreds of new proteins. This project addresses one of the critical steps in crystallography, the solution of the so-called 'phase problem.' Previous work within this project has resulted in the development of a powerful-direct methods algorithm, known as Shake-and-Bake, and a computer program, SnB, that can resolve the phase problem for structures with as many as 1300 unique non-hydrogen atoms. The selenium substructures of much larger proteins containing as many as 80 methionine residues can also be solved provided that these residues have been replaced by selenomethionine. The substructure can then be used as a bootstrap to locate the remainder of the protein molecule. The overall goal now is to extend the power and scope of the Shake-and- Bake algorithm. The ability of SnB to solve very large selenomethionine substructures (>100 Se sites) will be enhanced by the addition of a masking function to permit use of prior information concerning the molecular envelope or restrictions provided by non-crystallographic symmetry. Strategies will be devised for utilizing information from reference-beam diffraction to facilitate successful lower-resolution applications. Practical direct-methods algorithms will be developed for getting the most from diffraction data from one derivative (SIR) or for one wavelength with anomalous scattering (SAS). Concepts and routines from the PHASES program package will be used to integrate direct methods with other information and techniques commonly used in protein crystallography and to create pathway for the automated production of interpretable protein electron-density maps. Finally, the SnB website, http://www.hwi.buffalo.edu/SnB/, will be maintained for program dissemination and user education.