A versatile approach to high-density microcrystals in lipidic cubic phase for room-temperature serial crystallography.
A versatile approach to high-density microcrystals in lipidic cubic phase for room-temperature serial crystallography.
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DOI：
10.1107/s1600576723006428
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发表时间：
2023-10-01
影响因子：
6.1
通讯作者：
Moraes, Isabel
中科院分区：
文献类型：
--
作者：
Birch, James;Kwan, Tristan O. C.;Judge, Peter J.;Axford, Danny;Aller, Pierre;Butryn, Agata;Reis, Rosana I.;Juarez, Juan F. Bada;Vinals, Javier;Owen, Robin L.;Nango, Eriko;Tanaka, Rie;Tono, Kensuke;Joti, Yasumasa;Tanaka, Tomoyuki;Owada, Shigeki;Sugahara, Michihiro;Iwata, So;Orville, Allen M.;Watts, Anthony;Moraes, Isabel
关键词：
本文介绍了一种用于系列晶体学的高密度立方相膜蛋白微晶的制备方法。连续晶体学已成为膜蛋白结构研究的重要工具。在室温下以最小的辐射损伤从微米大小的弱衍射晶体中收集数据的能力为时间分辨研究和药物发现开辟了许多新的机会。然而，以所需的晶体密度和数量在无规立方相中生产完整的膜蛋白微晶是具有挑战性的。本文介绍了VIALS（多功能的方法，以高密度微晶的立方相连续晶体学），一个简单，快速和有效的方法，制备数百微升的高密度微晶适合在同步辐射和自由电子激光源的系列X射线衍射实验。该方法也是非常有益的合理的结构为基础的药物设计，因为它有利于在原位晶体浸泡和快速确定许多共晶结构。使用VIALS方法，室温结构报告（i）古视紫红质-3蛋白在其暗适应状态和110 ns光循环中间体，分别测定为2.2和1.7 μ m，和（ii）人A2 A腺苷受体与两种不同配体的复合物测定为3.5 μ m的分辨率。
A versatile approach for the preparation of high-density membrane protein microcrystals in lipidic cubic phase for serial crystallography is described. Serial crystallography has emerged as an important tool for structural studies of integral membrane proteins. The ability to collect data from micrometre-sized weakly diffracting crystals at room temperature with minimal radiation damage has opened many new opportunities in time-resolved studies and drug discovery. However, the production of integral membrane protein microcrystals in lipidic cubic phase at the desired crystal density and quantity is challenging. This paper introduces VIALS (versatile approach to high-density microcrystals in lipidic cubic phase for serial crystallography), a simple, fast and efficient method for preparing hundreds of microlitres of high-density microcrystals suitable for serial X-ray diffraction experiments at both synchrotron and free-electron laser sources. The method is also of great benefit for rational structure-based drug design as it facilitates in situ crystal soaking and rapid determination of many co-crystal structures. Using the VIALS approach, room-temperature structures are reported of (i) the archaerhodopsin-3 protein in its dark-adapted state and 110 ns photocycle intermediate, determined to 2.2 and 1.7 Å, respectively, and (ii) the human A2A adenosine receptor in complex with two different ligands determined to a resolution of 3.5 Å.
影响因子：
16.6
作者：
Bada Juarez JF;Judge PJ;Adam S;Axford D;Vinals J;Birch J;Kwan TOC;Hoi KK;Yen HY;Vial A;Milhiet PE;Robinson CV;Schapiro I;Moraes I;Watts A
通讯作者：
Watts A