Cryo-EM and artificial intelligence visualize endogenous protein community members

Ioannis Skalidis,Fotis L Kyrilis,Christian Tüting,Farzad Hamdi,Grzegorz Chojnowski,Panagiotis L Kastritis

doi:10.1016/j.str.2022.01.001

Ioannis Skalidis, Fotis L Kyrilis + Show 4 more

Open Access

https://doi.org/10.1016/j.str.2022.01.001

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Abstract

Cellular function is underlined by megadalton assemblies organizing in proximity, forming communities. Metabolons are protein communities involving metabolic pathways such as protein, fatty acid, and thioesters of coenzyme-A synthesis. Metabolons are highly heterogeneous due to their function, making their analysis particularly challenging. Here, we simultaneously characterize metabolon-embedded architectures of a 60S pre-ribosome, fatty acid synthase, and pyruvate/oxoglutarate dehydrogenase complex E2 cores de novo. Cryo-electron microscopy (cryo-EM) 3D reconstructions are resolved at 3.84-4.52 Å resolution by collecting <3,000 micrographs of a single cellular fraction. After combining cryo-EM with artificial intelligence-based atomic modeling and de novo sequence identification methods, at this resolution range, polypeptide hydrogen bonding patterns are discernible. Residing molecular components resemble their purified counterparts from other eukaryotes but also exhibit substantial conformational variation with potential functional implications. Our results propose an integrated tool, boosted by machine learning, that opens doors for structural systems biology spearheaded by cryo-EM characterization of native cell extracts.

Highlights

The cell is a very complex environment
Cryo-electron microscopy 3D reconstructions are resolved at 3.84–4.52 Aresolution by collecting
Our results propose an integrated tool, boosted by machine learning, that opens doors for structural systems biology spearheaded by Cryo-electron microscopy (cryo-EM) characterization of native cell extracts

Summary

Introduction

The cell is a very complex environment. Multiple processes have to be simultaneously coordinated and carried out in order for the cell to survive, perform its functions, and propagate. Community/metabolon-embedded proteins acquire different conformations to achieve function (Kyrilis et al, 2021b), and they can sometimes vastly differ in composition, structure, and function compared with studying them in isolation (Tu€ting et al, 2021). This is in contrast to their overexpressed and highly purified counterparts that are always preferred for structural studies due to their purity, homogeneity, and high quantity. The goal of modern structural biology should be to complement in vitro unambiguous data with the study of protein communities to more deeply understand endogenous cellular structure and function

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