Single‐particle cryo‐EM—Improved <i>ab initio</i> 3D reconstruction with SIMPLE/PRIME

TLDR

Cryogenic electron microscopy (cryo‑EM) and single‑particle analysis now enable high‑resolution structures of macromolecular assemblies that have resisted X‑ray crystallography and other approaches. The authors developed and extended the open‑source SIMPLE suite to analyze cryo‑EM single‑particle images, adding stochastic optimization algorithms to improve robustness. SIMPLE employs the probabilistic PRIME algorithm to cluster 2D images and determine 3D orientations, with added stochastic optimization to enhance robustness. The refined method yields ab initio 3D reconstructions with resolutions better than 10 Å, improves cluster center accuracy, and runs efficiently on standard laptops and workstations.

Abstract

Cryogenic electron microscopy (cryo-EM) and single-particle analysis now enables the determination of high-resolution structures of macromolecular assemblies that have resisted X-ray crystallography and other approaches. We developed the SIMPLE open-source image-processing suite for analysing cryo-EM images of single-particles. A core component of SIMPLE is the probabilistic PRIME algorithm for identifying clusters of images in 2D and determine relative orientations of single-particle projections in 3D. Here, we extend our previous work on PRIME and introduce new stochastic optimization algorithms that improve the robustness of the approach. Our refined method for identification of homogeneous subsets of images in accurate register substantially improves the resolution of the cluster centers and of the ab initio 3D reconstructions derived from them. We now obtain maps with a resolution better than 10 Å by exclusively processing cluster centers. Excellent parallel code performance on over-the-counter laptops and CPU workstations is demonstrated.