Eliminating the missing cone challenge through innovative approaches

IF 3.5 Q2 BIOCHEMISTRY & MOLECULAR BIOLOGY
Cody Gillman , Guanhong Bu , Emma Danelius , Johan Hattne , Brent L. Nannenga , Tamir Gonen
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Abstract

Microcrystal electron diffraction (MicroED) has emerged as a powerful technique for unraveling molecular structures from microcrystals too small for X-ray diffraction. However, a significant hurdle arises with plate-like crystals that consistently orient themselves flat on the electron microscopy grid. If the normal of the plate correlates with the axes of the crystal lattice, the crystal orientations accessible for measurement are restricted because the crystal cannot be arbitrarily rotated. This limits the information that can be acquired, resulting in a missing cone of information. We recently introduced a novel crystallization strategy called suspended drop crystallization and proposed that crystals in a suspended drop could effectively address the challenge of preferred crystal orientation. Here we demonstrate the success of the suspended drop approach in eliminating the missing cone in two samples that crystallize as thin plates: bovine liver catalase and the SARS‑CoV‑2 main protease (Mpro). This innovative solution proves indispensable for crystals exhibiting systematic preferred orientations, unlocking new possibilities for structure determination by MicroED.

Abstract Image

通过创新方法消除缺锥难题
微晶电子衍射(MicroED)已成为一种强大的技术,可用于揭示因晶体太小而无法进行 X 射线衍射的分子结构。然而,板状晶体在电子显微镜网格上始终保持平直方向时,会遇到一个重大障碍。如果平板的法线与晶格的轴线相关联,则可用于测量的晶体取向就会受到限制,因为晶体不能任意旋转。这就限制了可获取的信息,导致信息锥缺失。我们最近推出了一种名为悬滴结晶的新型结晶策略,并提出悬滴中的晶体可以有效解决首选晶体取向的难题。在这里,我们展示了悬滴法在两种结晶为薄板的样品(牛肝过氧化氢酶和 SARS-CoV-2 主要蛋白酶 (Mpro))中消除缺失锥的成功案例。事实证明,这种创新解决方案对于表现出系统性优选取向的晶体是不可或缺的,从而为通过 MicroED 进行结构测定开辟了新的可能性。
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来源期刊
Journal of Structural Biology: X
Journal of Structural Biology: X Biochemistry, Genetics and Molecular Biology-Structural Biology
CiteScore
6.50
自引率
0.00%
发文量
20
审稿时长
62 days
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