On the Feasibility of Clinical Studies with Cross-Linking Mass Spectrometry

In living systems, protein function relies on many intra- and intermolecular interactions within a network called the interactome. The majority of available interactome data has been acquired with isolated proteins and complexes, but visualization of interactome changes in living systems is crucial to advance understanding of functional changes with diseases and for the development of improved therapies. With model animal systems, quantitative cross-linking mass spectrometry has been successfully applied to uniquely reveal interactome changes with mitochondrial dysfunction both in heart failure and with age-related muscle function decline. In this study, we investigated the feasibility of qualitative cross-linking mass spectrometry for mitochondrial interactome studies with clinically relevant human muscle biopsy samples and amounts. Analysis of biopsy samples from two volunteers resulted in the identification of 1350 nonredundant peptides from 177 mitochondrial proteins from all mitochondrial subcompartments. Many of the identified human biopsy cross-linked peptides were derived from protein complex and supercomplex assemblies that exhibited altered levels in model systems of heart failure and aging. The findings demonstrate the initial feasibility that these and other cross-linked species can be detected in human muscle biopsy samples to enable future studies of age- and disease-related changes in mitochondrial structure–function relationships.