Electron counting and beam-induced motion correction enable near-atomic-resolution single-particle cryo-EM

Abstract

In recent work with large high symmetry viruses, single particle electron cryomicroscopy (cryoEM) has reached the milestone of determining near atomic resolution structures by allowing direct fitting of atomic models into experimental density maps. However, achieving this goal with smaller particles of lower symmetry remains extraordinarily challenging. Using a newly developed single electron counting detector, we confirm that electron beam induced motion significantly degrades resolution and, importantly, show how the combination of rapid readout and nearly noiseless electron counting allow image blurring to be corrected to subpixel accuracy. Thus, intrinsic image information can be restored to high resolution (Thon rings visible to ~3 Å). Using this approach we determined a 3.3 Å resolution structure of a ~700 kDa protein with D7 symmetry showing clear side chain density. Our method greatly enhances image quality and data acquisition efficiency - key bottlenecks in applying near atomic resolution cryoEM to a broad range of protein samples.