Equipment for X-ray Analysis of Biological Macromolecules

这项提案要求提供资金，以升级德克萨斯大学使用X射线衍射进行生物大分子分析的设施。该设备将用于两名现有研究人员和两名新任命的助理教授的研究。它分为三类：A.X射线衍射、B.高速计算和C.分子图形学。答：衍射设备由高亮度旋转阳极X射线发生器和最先进的成像板(IP)区域探测器组成。探测器捕获X射线，作为BaFBr：eux荧光粉上的亚稳定中心，随后通过激光激活读取该荧光粉。这种类型的探测器具有空间分辨率，既可以容纳单晶蛋白质衍射，也可以容纳设备用户产生的低角散射图案。B.一台高速、双处理器、磁盘容量为4 GB的计算机将大量用于分子模型的精细化。这台机器将允许使用程序XploR及时周转模拟退火精细化。它还将用于对低角散射模式的复杂分析。C.将获得一个现代分子图像站。这台仪器将增加大量使用的分子图形容量，并可能需要更换目前正在使用的故障仪器。用户群体的研究兴趣相当不同，涉及许多生物学上重要的系统，正在用一系列方法进行研究。通过单晶分析研究的蛋白质包括：核糖体抑制蛋白，如蓖麻毒素、PAP和eBulin；植物和真菌几丁质酶；抗真菌蛋白质Zeamatin；至少四种结构不同的氨基酸脱羧酶，包括鸟氨酸、精氨酸和组氨酸脱羧酶；几种无脊椎动物血红蛋白；多酶复合体，如PDC；以及核糖体RNA结合蛋白L9。除单晶研究外，该装置还将用于可聚合脂质组装体的低角散射分析。目前正在进行系统的研究，以确定管理脂质双层平面内的亚组装体的规则。上述项目往往远远超出了对初始X射线结构的简单描述。每一种蛋白质在结晶学意义上都被提炼到其数据的极限，以便产生尽可能准确的分子模型。此外，在适当的情况下对底物类似物或其他配体的结合进行了评估。几个研究小组都有强有力的定点突变计划，这些计划创造了一个蛋白质家族，其X射线分析对于描述它们的结构/功能关系很重要。最后，基于结构的药物设计正在开发中。设计范式需要对众多中间结构进行分析和能量精炼。所要求的设备对于现有方案的继续和新方案的扩展非常重要。X射线发生器和激电探测器以及冷温设备对于正在研究的许多系统的数据收集都很重要。其中一些晶体系统有很大的晶胞，需要很长的时间或几个晶体才能完成数据收集。一些项目涉及大量突变、抑制或连接的结构，在所有情况下，都希望收集尽可能高分辨率的数据。所有这些考虑都要求提高数据收集能力。此外，由于技术原因，在没有新设备的情况下，不能开展涉及双层结构的低角度散射工作的研究项目。所要求的计算升级将允许XploR等现代结构优化程序快速运行。它还将使用于单晶和低角度散射分析的最现代和最强大的数据分析程序相对容易地执行。大学管理的超级计算机的明显消亡加剧了对这个系统的需求。由于有十几名学生和其他科学家在使用这些设备，目前的分子图形设备存在压力。一个新的单位将允许对正在进行的大量结构工作进行适当的分析，特别是在目前到位的一个关键仪器发生故障的情况下。

This proposal requests funds to upgrade facilities for analysis of biological macromolecules, using X-ray diffraction, at the University of Texas. The equipment will be used in the research of two established investigators and two newly appointed assistant professors. It falls into three categories, A. X-ray diffraction, B. high speed computing, and C. molecular graphics. A. The diffraction equipment consists of a high brilliance rotating anode X-ray generator and a state of the art imaging plate (IP) area detector. The detector captures X-rays as metastable centers on a BaFBr:EuX-phosphor, which is read by subsequent laser activation. This type of detector has the spatial resolution to accommodate both single crystal protein diffraction and the low angle scattering patterns generated by the users of the equipment. B. A high speed, dual processor, computer with 4 Gbytes of disk capacity will be used largely for molecular model refinement. This machine will allow timely turnover of simulated annealing refinements using the program XPLOR. It will also be used for sophisticated analysis of low angle scattering patterns. C. A modern molecular graphics station will be acquired. This instrument will augment a heavily used molecular graphics capacity and may be required to replace a failing instrument currently in use. The research interests of the user group are quite varied, involving a number of biologically important systems under study by a range of methodologies. Proteins under study by single crystal analysis include: ribosome inhibiting proteins like ricin, PAP and ebulin; plant and fungal chitinases; the antifungal protein zeamatin; at least four structurally different amino acid decarboxylases including ornithine, arginine and histidine decarboxylases; several invertebrate hemoglobins; multienzyme complexes, like PDC; and the ribosomal RNA binding protein L9. In addition to single crystal studies, the equipment will be used in the low angle scattering ana lysis of polymerizable lipid assemblies. Systematic studies are underway to define the rules governing subassembly in the planes of lipid bilayers. The above projects tend to extend well beyond simply describing the initial X-ray structure. Each protein is refined in a crystallographic sense to the limits of it data in order to produce as accupate a molecular model as possible. Also, the binding of substrate analogs or other ligands is assessed where appropriate. Several of the research groups have vigorous site-directed mutagenesis programs which create a family of proteins whose X-ray analysis is important to describing their structure/function relationships. Finally, structure-based drug design is being developed. The design paradigm requires analysis and energy refinement of numerous intermediate structures. The equipment requested is of great importance to the continuation of established programs and to the expansion of new ones. The X-ray generator and IP detector, together with a cold temperature apparatus, are important for data collection on the many systems under study. Some of these crystal systems have large unit cells and require lengthy time periods or several crystals for complete data collection. Some projects involve a large number of mutant, inhibited, or liganded structures, and in all cases it is desirable to collect as high resolution data as possible. All of these considerations argue for increased data collecting capacity. Furthermore, research projects involving low angle scattering work on bilayer structures cannot be carried out, for technical reasons, without acquisition of the new equipment. The requested computing upgrade will allow modern structural refinement programs, such as XPLOR, to run expeditiously. It will also allow the most modern and powerful data analysis programs for single crystal and low angle scattering analysis to be executed with comparative ease. The need for this system is intensified by the apparent demise of the University managed supercomputer. With over a dozen students and other scientists using the facilities, there is a strain on current molecular graphics equipment. A new unit will allow proper analysis of the mass of structural work being done particularly if one key instrument, currently in place, fails.