Characterization and Crystallization of Pathogenic Protozoa Proteins

致病性原生动物医学结构基因组学（MSGPP）项目的总体目标是确定具有小分子配体复合物的致病性原生动物的生物医学重要蛋白质的晶体结构，并利用该信息加速药物开发，以治疗和预防由10种主要致病性生物引起的疾病。MSGPP正处于基因组测序和结构基因组学革命的十字路口，将核苷酸序列数据转化为蛋白质、晶体、三维结构和蛋白质抑制化合物。Project 3对于这个紧密结合的Program Project至关重要，因为它位于Project 1和Projects的交汇处，Project 1将提供配体，Project 1将提供大量的MSGPP蛋白质样品，而Project 4的主要产出将是用于x射线结构测定的蛋白质-配体复合物晶体。我们将使用多种途径获得x射线衍射质量晶体。在第一步中，将执行几个表征步骤，以了解如何修剪蛋白质，用配体稳定，提供低多分散诱导缓冲液，并进行化学修饰。在第二步中，一系列互补的方法，特别是一套集成的机器人和数据库，将与几个重点优化方法一起应用于晶体生长。在高通量初始晶体筛选、二次维里系数测定、毛细管等方面开展了创新合作

The overall goal of the Medical Structural Genomics of Pathogenic Protozoa (MSGPP) Program Project is to determine crystal structures of biomedically important proteins of pathogenic protozoa in complex with small-molecule ligands, and to use that information for accelerated drug development for treatment and prevention of the diseases caused by the ten major pathogenic organisms targeted. MSGPP is on the crossroads of the genomes sequencing and structural genomics revolutions in translating nucleotide sequence data into proteins, crystals, three-dimensional structures and protein-inhibiting compounds. This Project 3 is critical for this tightly integrated Program Project since it is at the intersection of Project 1, which will provide Ligands, and Projects, which will provide numerous MSGPP protein samples, while the major output of this Project will be crystals if protein-ligand complexes for X-ray structure determination in Project 4. We will use multiple avenues to obtain X-ray diffraction quality crystals. In a first step, several characterization steps will be performed to see how the protein can be trimmed, stabilized with ligands, provided with a low-polydispersity inducing buffer, and chemically modified. In a second step, a series of complementary approaches and in particular an integrated set of robots and databases will be applied to grow crystals in conjunction with several focused optimization methodologies. Innovative collaborations are in place for high-throughput initial crystal screening, determination of second virial coefficients, capillary crystallization robotics, and automated crystal image analysis. The entire spectrum of approaches within MSGPP is estimated to result in an overall crystallization success rate between 30 to 40% for each group of target variants.