The fast development of single-particle cryogenic electron microscopy (cryo-EM) has made it more feasible to obtain the 3D structure of well-behaved macromolecules with a molecular weight higher than 300 kDa at ~3 Å resolution. However, it remains a challenge to obtain the high-resolution structures of molecules smaller than 200 kDa using single-particle cryo-EM. In this work, we apply the Cs-corrector-VPP-coupled cryo-EM to study the 52 kDa streptavidin (SA) protein supported on a thin layer of graphene and embedded in vitreous ice. We are able to solve both the apo-SA and biotin-bound SA structures at near-atomic resolution using single-particle cryo-EM. We demonstrate that the method has the potential to determine the structures of molecules as small as 39 kDa.

单粒子低温电子显微镜（cryo-EM）的快速发展使以3Å分辨率获得分子量高于300 kDa的行为良好的大分子的3D结构变得更加可行。然而，使用单粒子冷冻EM获得小于200 kDa的分子的高分辨率结构仍然是一个挑战。在这项工作中，我们应用Cs-校正剂-VPP耦合的冷冻电磁场研究了支撑在石墨烯薄层上并嵌入玻璃冰中的52 kDa链霉亲和素（SA）蛋白。我们能够使用单粒子冷冻电磁场以接近原子的分辨率解决apo-SA和生物素结合的SA结构。我们证明该方法具有确定小至39 kDa分子结构的潜力。