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首页> 外文期刊>Acta crystallographica.Section D. Biological crystallography >High-resolution neutron protein crystallography with radically small crystal volumes: application of perdeuteration to human aldose reductase
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High-resolution neutron protein crystallography with radically small crystal volumes: application of perdeuteration to human aldose reductase

机译：高分辨率中子蛋白质晶体学与彻底的小水晶卷:应用程序perdeuteration人类醛糖还原酶

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Neutron diffraction data have been collected to 2.2 angstrom resolution from a small (0.15 mm(3)) crystal of perdeuterated human aldose reductase (h-AR; MW=36 kDa) in order to help to determine the protonation state of the enzyme. h-AR belongs to the aldo-keto reductase family and is implicated in diabetic complications. Its ternary complexes (h-AR-coenzyme NADPH-selected inhibitor) provide a good model to study both the enzymatic mechanism and inhibition. Here, the successful production of fully deuterated human aldose reductase [h-AR(D)], subsequent crystallization of the ternary complex h-AR(D)-NADPH-IDD594 and neutron Laue data collection at the LADI instrument at ILL using a crystal volume of just 0.15 mm(3) are reported. Neutron data were recorded to 2 angstrom resolution, with subsequent data analysis using data to 2.2 angstrom. This is the first fully deuterated enzyme of this size (36 kDa) to be solved by neutron diffraction and represents a milestone in the field, as the crystal volume is at least one order of magnitude smaller than those usually required for other high-resolution neutron structures determined to date. This illustrates the significant increase in the signal-to-noise ratio of data collected from perdeuterated crystals and demonstrates that good-quality neutron data can now be collected from more typical protein crystal volumes. Indeed, the signal-to-noise ratio is then dominated by other sources of instrument background, the nature of which is under investigation. This is important for the design of future instruments, which should take maximum advantage of the reduction in the intrinsic diffraction pattern background from fully deuterated samples.
机译：中子衍射数据收集2.2埃分辨率从一个小(0.15毫米(3))水晶perdeuterated人类的醛糖还原酶(h-AR;质子化作用的酶。aldo-keto还原酶家族,与糖尿病并发症。配合物(h-AR-coenzyme NADPH-selected抑制剂)提供一个良好的模型来研究的酶和抑制机制。成功的生产完全氘人类醛糖还原酶(h-AR (D)),随后三元复杂的结晶h-AR (D) -NADPH-IDD594和中子劳厄数据收集在LADI仪器使用晶体的体积只有0.15毫米(3)报告。中子数据记录2埃解决,后续数据分析使用数据为2.2埃。这种规模的氘酶(36 kDa)通过中子衍射和解决该领域的里程碑,因为晶体体积至少一个数量级小于这些通常需要其他高分辨率中子结构决定日期。说明了显著增加收集的数据的信噪比perdeuterated晶体和证明高质量的中子数据现在可以收集更典型的蛋白质晶体卷。事实上,信噪比是由其他来源的乐器背景下的性质调查。未来的工具,应该最大减少内在的优势从完全衍射模式的背景氘样本。

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