Post-translational modifications of proteins in cardiovascular diseases examined by proteomic approaches

Abstract

Over 400 different types of post-translational modifications (PTMs) have been reported and over 200 various types of PTMs have been discovered using mass spectrometry (MS)-based proteomics. MS-based proteomics has proven to be a powerful method capable of global PTM mapping with the identification of modified proteins/peptides, the localization of PTM sites and PTM quantitation. PTMs play regulatory roles in protein functions, activities and interactions in various heart related diseases, such as ischemia/reperfusion injury, cardiomyopathy and heart failure. The recognition of PTMs that are specific to cardiovascular pathology and the clarification of the mechanisms underlying these PTMs at molecular levels are crucial for discovery of novel biomarkers and application in a clinical setting. With sensitive MS instrumentation and novel biostatistical methods for precise processing of the data, low-abundance PTMs can be successfully detected and the beneficial or unfavorable effects of specific PTMs on cardiac function can be determined. Moreover, computational proteomic strategies that can predict PTM sites based on MS data have gained an increasing interest and can contribute to characterization of PTM profiles in cardiovascular disorders. More recently, machine learning- and deep learning-based methods have been employed to predict the locations of PTMs and explore PTM crosstalk. In this review article, the types of PTMs are briefly overviewed, approaches for PTM identification/quantitation in MS-based proteomics are discussed and recently published proteomic studies on PTMs associated with cardiovascular diseases are included.

Keywords: MS‐based proteomics; cardiovascular disease; post‐translational modifications; proteins.

Fig. 1

Schematically shown post‐translational modifications (PTMs) that are covered in PTMs relation to heart and cardiovascular diseases and their association with heart and cardiovascular diseases as identified by mass spectrometry (MS)‐based proteomics. Additional details can be found in the text.

Fig. 2

Schema of a clickable glutathione approach for identification of glutathionylated peptides/proteins by LC–MS/MS. (A) HL‐1 cells were transfected with a mutant of glutathione synthetase GSM4, which uses azido‐alanine to synthesize azido‐glutathione N₃‐GSH. After addition of reactive oxygen species (ROS) stimulus (H₂O₂), glutathionylated proteins containing azide groups were enriched using a click reaction with the alkyne functional group of biotin‐alkyne. (B) formula of biotin‐DADPS‐alkyne used in the present study for isolation and elution of glutathionylated peptides. (C) After the click reaction, biotinylated glutathionylated proteins were bound into streptavidin‐agarose beads, then on‐bead digested by trypsin/Lys‐C, resulting in glutathionylated peptides eluted by acidic cleavage of DADPS linker and identified by LC–MS/MS. Additional details are provided in the text. Figure adapted from VanHecke et al. [33]; copyright (2019) American Chemical Society.

Fig. 3

The structure of sarcomere, the basic contractile unit of the heart, showing the proteins and their altered phosphorylation that were identified in ischemic cardiomyopathy (ICM) patients. The sarcomere contains thin actin‐based filament and thick myosin‐based filament laterally bordered by the Z‐disk. Decreased expression and phosphorylation levels of cardiac troponin I (cTnI) and enigma homolog 2 (ENH2) were detected, as well as increased phosphorylation of muscle LIM protein (MLP) and calsarcin‐1 (Cal‐1) in ICM cardiac tissues. Other proteins depicted: cysteine‐rich protein 2 (CRIP2), cardiac troponin T (cTnT), tropomyosin (Tpm), myosin light chains (MLCs) and troponin C (TnC). Figure adapted from Chapman et al. [1]; copyright (2023) American Chemical Society.

Fig. 4

The structural model and sequence of β‐myosin heavy chain (β‐MHC) with newly identified post‐translational modification (PTM) sites. (A) X‐ray crystal structure of myosin motor domain; the four motor subdomains [N‐terminal domain (yellow), upper 50 kDa domain (cyan), lower 50 kDa domain (light blue) and converter domain (dark blue)] are shown with identified PTMs (acetylation and phosphorylation) in red colors (K213‐Ac, K429‐Ac, K58‐Ac, K34‐Ac, S210‐P and T215‐P); the inset highlights the nucleotide binding pocket and functional loops (loop 1, switches 1 and 2, and phosphate binding loop). (B) Schema of β‐MHC sequence with key functional regions and locations of PTMs; colored blocks correspond to motor subdomains shown in (A). (C) X‐ray crystal structure of S2 region of myosin coiled‐coil tail with K951‐Ac site depicted (blue, chain A; green, chain B). Figure adapted from Landim‐Vieira et al. [136].