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An informatic framework for decoding protein complexes by top-down mass spectrometry.

ABSTRACT: Efforts to map the human protein interactome have resulted in information about thousands of multi-protein assemblies housed in public repositories, but the molecular characterization and stoichiometry of their protein subunits remains largely unknown. Here, we report a computational search strategy that supports hierarchical top-down analysis for precise identification and scoring of multi-proteoform complexes by native mass spectrometry.

SUBMITTER: Skinner OS

PROVIDER: S-EPMC4767540 | biostudies-literature | 2016 Mar

REPOSITORIES: biostudies-literature

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An informatic framework for decoding protein complexes by top-down mass spectrometry.

Skinner Owen S OS Havugimana Pierre C PC Haverland Nicole A NA Fornelli Luca L Early Bryan P BP Greer Joseph B JB Fellers Ryan T RT Durbin Kenneth R KR Do Vale Luis H F LH Melani Rafael D RD Seckler Henrique S HS Nelp Micah T MT Belov Mikhail E ME Horning Stevan R SR Makarov Alexander A AA LeDuc Richard D RD Bandarian Vahe V Compton Philip D PD Kelleher Neil L NL

Nature methods 20160118 3

Efforts to map the human protein interactome have resulted in information about thousands of multi-protein assemblies housed in public repositories, but the molecular characterization and stoichiometry of their protein subunits remains largely unknown. Here, we report a computational search strategy that supports hierarchical top-down analysis for precise identification and scoring of multi-proteoform complexes by native mass spectrometry. ...[more]

PMID: 26780093

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Project description:Mass spectrometry (MS)-based proteomics is playing an increasingly important role in cardiovascular research. Proteomics includes identification and quantification of proteins and the characterization of protein modifications, such as posttranslational modifications and sequence variants. The conventional bottom-up approach, involving proteolytic digestion of proteins into small peptides before MS analysis, is routinely used for protein identification and quantification with high throughput and automation. Nevertheless, it has limitations in the analysis of protein modifications, mainly because of the partial sequence coverage and loss of connections among modifications on disparate portions of a protein. An alternative approach, top-down MS, has emerged as a powerful tool for the analysis of protein modifications. The top-down approach analyzes whole proteins directly, providing a "bird's-eye" view of all existing modifications. Subsequently, each modified protein form can be isolated and fragmented in the mass spectrometer to locate the modification site. The incorporation of the nonergodic dissociation methods, such as electron-capture dissociation (ECD), greatly enhances the top-down capabilities. ECD is especially useful for mapping labile posttranslational modifications that are well preserved during the ECD fragmentation process. Top-down MS with ECD has been successfully applied to cardiovascular research, with the unique advantages in unraveling the molecular complexity, quantifying modified protein forms, complete mapping of modifications with full-sequence coverage, discovering unexpected modifications, identifying and quantifying positional isomers, and determining the order of multiple modifications. Nevertheless, top-down MS still needs to overcome some technical challenges to realize its full potential. Herein, we reviewed the advantages and challenges of the top-down method, with a focus on its application in cardiovascular research.

Dataset Information

An informatic framework for decoding protein complexes by top-down mass spectrometry.

Publications

An informatic framework for decoding protein complexes by top-down mass spectrometry.

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