Top-down Proteomics: Technology Advancements and Applications to Heart Diseases

Abstract

Introduction Diseases of the heart are a leading cause of morbidity and mortality for both men and women worldwide, and impose significant economic burdens on the healthcare systems. Despite substantial effort over the last several decades, the molecular mechanisms underlying diseases of the heart remain poorly understood. Areas covered Altered protein post-translational modifications (PTMs) and protein isoform switching are increasingly recognized as important disease mechanisms. Top-down high-resolution mass spectrometry (MS)-based proteomics has emerged as the most powerful method for the comprehensive analysis of PTMs and protein isoforms. Here, we will review recent technology developments in the field of top-down proteomics, as well as highlight recent studies utilizing top-down proteomics to decipher the cardiac proteome for the understanding of the molecular mechanisms underlying diseases of the heart. Expert commentary Top-down proteomics is a premier method for the global and comprehensive study of protein isoforms and their PTMs, enabling the identification of novel protein isoforms and PTMs, characterization of sequence variations, and quantification of disease-associated alterations. Despite significant challenges, continuous development of top-down proteomics technology will greatly aid the dissection of the molecular mechanisms underlying diseases of the hearts for the identification of novel biomarkers and therapeutic targets.

Role of myosin light chain phosphatase in cardiac physiology and pathophysiology

“…Two-dimensional electrophoresis with isoelectric focusing can also be used to separate nonphosphorylated and monophosphorylated RLC, but it is a more laborious procedure [81]. Additionally, more sophisticated and detailed analyses of heart extracts utilizing top-down approaches of mass spectrometry have been described which provide measurements of various post-translational modifications of RLC and other sarcomeric proteins [22, 87]. …”

Identification and Quantification of Proteoforms by Mass Spectrometry

“…Top‐down proteomics directly analyzes intact proteins . The relationship between amino acid sequence and PTMs is preserved and proteoforms can be characterized, providing a proteoform‐specific understanding of biological phenomena.…”

The Impact of Phosphorylation on Electron Capture Dissociation of Proteins: A Top-Down Perspective

“…Since the introduction of electron capture dissociation (ECD) in 1998 [1], this powerful tandem mass spectrometry (MS/MS) technique has been widely used to elucidate amino acid sequences, structures, and, particularly, to localize post-translational modifications (PTMs) of proteins [2–6]. During ECD, ion analytes (proteins or peptides) are irradiated by low-energy electrons, cleaving N-C α bonds and producing c -type and z -type ions [1, 2, 7, 8].…”

“…Phosphorylation, a highly prevalent PTM, plays a critical role in regulating various complex cellular processes, and altered phosphorylation is implicated in many diseases [5, 6, 18]. During CAD, the phosphorylated ions often experience 98 and 80 Da neutral losses, hindering the interrogation of phosphorylation sites [19].…”