Complexome Profiling: Assembly and Remodeling of Protein Complexes.

Ilka Wittig,Pedro Felipe Malacarne

doi:10.3390/ijms22157809

Ilka Wittig, Pedro Felipe Malacarne

Open Access

https://doi.org/10.3390/ijms22157809

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Abstract

Many proteins have been found to operate in a complex with various biomolecules such as proteins, nucleic acids, carbohydrates, or lipids. Protein complexes can be transient, stable or dynamic and their association is controlled under variable cellular conditions. Complexome profiling is a recently developed mass spectrometry-based method that combines mild separation techniques, native gel electrophoresis, and density gradient centrifugation with quantitative mass spectrometry to generate inventories of protein assemblies within a cell or subcellular fraction. This review summarizes applications of complexome profiling with respect to assembly ranging from single subunits to large macromolecular complexes, as well as their stability, and remodeling in health and disease.

Highlights

The function of many proteins often requires stable or dynamic associations with other biomolecules, e.g., proteins, nucleic acids, carbohydrates or lipids in order to form large macromolecular assemblies
Targeted interaction proteomics may involve affinity enrichment protocols that rely on antibodies [6,7], affinity tags [8,9,10], or in cell biotin labeling [11] coupled to quantitative mass spectrometry
The advantage of using targeted strategies is an enrichment of the protein assembly, which enables in depth characterization of its interacting components

Summary

Introduction

The function of many proteins often requires stable or dynamic associations with other biomolecules, e.g., proteins, nucleic acids, carbohydrates or lipids in order to form large macromolecular assemblies. Very useful for studies on large assemblies of biomolecules, density gradients have inherent low resolution, require a large amount of sample, and it is difficult to differentiate between co-purification in a fraction or an actual physical protein– protein interaction. BNE became a very robust and reproducible, micro-scale high resolution separation method to examine composition of protein complexes in a broad range of samples from bacterial membranes, soluble subcellular components, and membrane fractions from eukaryotic cells to tissue specimens from patients [34]. The aim of each complexome profiling experiment is to gain information of native protein complexes and to compare the appearance and abundance of assemblies under different conditions or in disease states. Further developments on CP (Table 1) included steps in the sample preparation discussed in the following sub-sections; (1) to enable complex assembly to be monitored, (2) to improve the sample comparison using SILAC and TMT, to monitor protein complex, (3) remodeling, (4) turnover and repair, (5) to gain structural information on complex conformations by applying protein crosslinkers, and (6) to identify RNAprotein complexes

Assembly and Stability of OXPHOS Complexes

Multiplexing CP

Remodeling

Turnover of Subunits within Protein Complexes

Structural Information of Protein Complex Conformation

RNA-Protein Complexes

Data Analysis

Conclusions