Developing Structural Interactomics and its Application in Cell Biology

Abstract

Proteins are involved in almost all cellular processes. They are organized through extensive networks of interactions, regulating the working mechanisms of the cell with high fidelity and precision. However, characterization of the protein interactome has been an extremely challenging task due to the lack of highly confident and efficient technology. To address this issue, we have developed a high‐throughput approach for comprehensive protein interactome profiling based on cross‐linking mass spectrometry (XL‐MS). We applied this novel XL‐MS strategy to several highly complex samples, including various cell lysates as well as organelles such as the endoplasmic reticulum the mitochondrion. In particular, in organelle XL‐MS allows covalent capturing of the entire interaction network of all membrane proteins in association with their regulatory partners. Thus, it captures a snapshot of functional protein networks in vivo prior to protein extraction under detergent conditions, enabling directly profiling of the native assembly of macromolecular machineries and their cross‐talk.We have demonstrated the diverse capabilities of XL‐MS in detail by our recent interactome study of mouse heart mitochondria. The obtained mitochondrial interaction network is comprehensive (more than three thousand cross‐links were identified, covering all major mitochondrial membrane protein complexes), confident (cross‐links are validated on existing high‐resolution structures) and specific (cross‐links are highly specific within each mitochondrial sub‐compartment). Remarkably, through thousands of protein‐protein interactions, we elucidated several organization principles of mitochondrial macromolecular assemblies, discovered previously unrecognized protein interactions, and revealed novel mitochondrial proteins.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.