DIFFRACTION OF BACTERIAL ENDOTOXINS: STRUCT BASED DRUG DESIGN, SCREEN PCP ENZYME

细菌内毒素的衍射：基于结构的药物设计，筛选 PCP 酶

• 批准号: 6667781
• 负责人: Vivian Cody
• 金额: $ 14.27万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-09-30 至 2003-08-14
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6667781
• 关键词: bioimaging /biomedical imaging; biological products; biomedical resource; proteins; radiography; structural biology

Delta endotoxins are the entomocidal agent produced by Bacillus thuringiensis ssp. Kurstaki. These toxins are produced as protoxins (135 kDa) by the bacteria during sporulation and recognized by their bipyrimidal crystalline form in the sporangium. When ingested by larval Lepidoptera the crystal dissolves and is proteolytically digested to the active toxin form (60-65 kDa). This activated toxin is capable of opening K+ channels in the epithelium of the insect midgut. Ion leakage ultimately results in the complete disruption of the midgut and death of the insect. More recently, a new active form of the toxin, CryIIIB2, has been determined (64 kDa). The bacterial endotoxin CryIIIB2 is a 652 residue protein which crystallizes in the orthorhombic space group C2221 with unit cell dimensions a = 122.44, b = 131.81, and c = 105.37& and contain one molecule in the asymmetric unit. Synchrontron radiation was used to collect 2.2& resolution data at the CHESS facility at Cornell University in December, 1997 by Vivian Cody and Nikolai Galitsky for the complex of EC11095 with D-galactosamine. Although these data were did not diffract uniformly to high resolution, Walt Pangborn was able to scale data with at least 50% completion for all shells to 2.2& resolution. The structure of the native CryIIIB2 was used as a search model for the rotation and translation function in the program XPLOR which revealed there was no significant change in the molecular orientation of the molecule compared to the room temperature model. The major consequence of low temperature data collection is the contraction of the unit cell along the a and b lattice directions. Interpretation of the difference electron density maps made from the 3.0& refinement model of the Arg-348 mutant of CryIIIB2 revealed a large density profile that clearly was not a cluster of solvent and that could be fit to the sugar. These data reveal that the galactosamine is positioned at the intersection of the three domains of CryIIIB2. Refinement of these data to 2.2& resolution suggest other potential sugar binding sites are possible.

δ内毒素是芽孢杆菌产生的杀虫剂 苏云金亚种 库斯塔基。 这些毒素以原毒素的形式产生 (135 kDa) 在孢子形成期间被细菌识别并被其识别 孢子囊中呈双嘧啶晶体形式。 当摄入时 幼虫鳞翅目晶体溶解并被蛋白水解 消化成活性毒素形式（60-65 kDa）。 这种激活的毒素 能够打开昆虫上皮细胞中的 K+ 通道 中肠。 离子泄漏最终导致完全破坏 昆虫的中肠和死亡。 最近，一种新的活跃形式 毒素 CryIIIB2 已被测定 (64 kDa)。 细菌 内毒素 CryIIIB2 是一种 652 个残基的蛋白质，在 正交空间群 C2221，晶胞尺寸 a = 122.44, b = 131.81，且 c = 105.37& 并且在不对称结构中包含一个分子 单元。 使用同步辐射来收集2.2&分辨率数据 1997 年 12 月在康奈尔大学国际象棋设施 Vivian Cody 和 Nikolai Galitsky 的 EC11095 复合物 D-半乳糖胺。 尽管这些数据并未均匀衍射 到高分辨率，Walt Pangborn 能够将数据扩展至至少 所有 shell 的完成度为 50%，达到 2.2& 分辨率。 的结构 原生 CryIIIB2 被用作旋转和搜索模型 XPLOR 程序中的翻译功能表明没有 分子的分子取向发生显着变化 与室温模型相比。 低的主要后果 温度数据收集是晶胞沿着 a 和 b 晶格方向。 差异的解释 由 3.0 和细化模型制作的电子密度图 CryIIIB2 的 Arg-348 突变体显示出大的密度分布， 显然不是溶剂簇，并且可以适合 糖。 这些数据表明，半乳糖胺位于 CryIIIB2 的三个结构域的交集。 对这些的细化 2.2&分辨率的数据表明其他潜在的糖结合位点 是可能的。