X-rays in the Cryo-Electron Microscopy Era: Structural Biology’s Dynamic Future

TLDR

Cryo‑electron microscopy has risen to rival X‑ray crystallography for near‑atomic resolution, yet crystallography still excels at small, highly ordered proteins and dynamic studies, while cryo‑EM excels at large, heterogeneous assemblies, and the future of structural biology depends on leveraging each method’s strengths. The authors aim to share expert insights and evaluate how X‑ray crystallography and cryo‑EM can jointly advance structural biology. They present a perspective that compares the unique and complementary capabilities of X‑ray crystallography and cryo‑EM to guide future research.

Abstract

Over the past several years, single-particle cryo-electron microscopy (cryo-EM) has emerged as a leading method for elucidating macromolecular structures at near-atomic resolution, rivaling even the established technique of X-ray crystallography. Cryo-EM is now able to probe proteins as small as hemoglobin (64 kDa) while avoiding the crystallization bottleneck entirely. The remarkable success of cryo-EM has called into question the continuing relevance of X-ray methods, particularly crystallography. To say that the future of structural biology is either cryo-EM or crystallography, however, would be misguided. Crystallography remains better suited to yield precise atomic coordinates of macromolecules under a few hundred kilodaltons in size, while the ability to probe larger, potentially more disordered assemblies is a distinct advantage of cryo-EM. Likewise, crystallography is better equipped to provide high-resolution dynamic information as a function of time, temperature, pressure, and other perturbations, whereas cryo-EM offers increasing insight into conformational and energy landscapes, particularly as algorithms to deconvolute conformational heterogeneity become more advanced. Ultimately, the future of both techniques depends on how their individual strengths are utilized to tackle questions at the frontiers of structural biology. Structure determination is just one piece of a much larger puzzle: a central challenge of modern structural biology is to relate structural information to biological function. In this perspective, we share insight from several leaders in the field and examine the unique and complementary ways in which X-ray methods and cryo-EM can shape the future of structural biology.