N-Terminal Proteomics Reveals Distinct Protein Degradation Patterns in Different Types of Human Atherosclerotic Plaques.

Atherosclerotic plaque rupture is a major cause of cardiovascular events. Plaque destabilization is associated with extracellular matrix (ECM) modification involving proteases which generate protein fragments with new N-termini. We hypothesized that rupture-prone plaques would contain elevated fragment levels, and their sequences would allow identification of active proteases and target proteins.

Proteomic analysis of the extracellular matrix of human atherosclerotic plaques shows marked changes between plaque types.

Cardiovascular disease is the leading cause of death, with atherosclerosis the major underlying cause. While often asymptomatic for decades, atherosclerotic plaque destabilization and rupture can arise suddenly and cause acute arterial occlusion or peripheral embolization resulting in myocardial infarction, stroke and lower limb ischaemia. As extracellular matrix (ECM) remodelling is associated with plaque instability, we hypothesized that the ECM composition would differ between plaques.

The inflammatory oxidant peroxynitrous acid modulates the structure and function of the recombinant human V3 isoform of the extracellular matrix proteoglycan versican.

Continued oxidant production during chronic inflammation generates host tissue damage, with this being associated with pathologies including atherosclerosis. Atherosclerotic plaques contain modified proteins that may contribute to disease development, including plaque rupture, the major cause of heart attacks and strokes. Versican, a large extracellular matrix (ECM) chondroitin-sulfate proteoglycan, accumulates during atherogenesis, where it interacts with other ECM proteins, receptors and hyaluronan, and promotes inflammation.

Identification of galectin-1 and other cellular targets of alpha,beta-unsaturated carbonyl compounds, including dimethylfumarate, by use of click-chemistry probes.

α,β-Unsaturated carbonyls are a common motif in environmental toxins (e.g. acrolein) as well as therapeutic drugs, including dimethylfumarate (DMFU) and monomethylfumarate (MMFU), which are used to treat multiple sclerosis and psoriasis.

Peroxynitrous acid-modified extracellular matrix alters gene and protein expression in human coronary artery smooth muscle cells and induces a pro-inflammatory phenotype.

Leukocytes produce oxidants at inflammatory sites, including within the artery wall during the development of atherosclerosis. Developing lesions contain high numbers of activated leukocytes that generate reactive nitrogen species, including peroxynitrite/peroxynitrous acid (ONOO/ONOOH), as evidenced by the presence of oxidized/nitrated molecules including extracellular matrix (ECM) proteins. ECM materials are critical for arterial wall integrity, function, and determine cell phenotype, with smooth muscle cells undergoing a phenotypic switch from quiescent/contractile to proliferative/synthetic during disease development.

Proteomic Characterization of Atherosclerotic Lesions In Situ Using Percutaneous Coronary Intervention Angioplasty Balloons-Brief Report.

Background: Materials extracted from atherosclerotic arteries can disclose data about the molecular pathology of cardiovascular disease, but obtaining such samples is complex and requires invasive surgery. To overcome this barrier, this study investigated whether angioplasty balloons inflated during standard percutaneous coronary interventions retain proteins from treated (dilated) atherosclerotic lesions and whether proteomic analysis of this material could provide data on lesion protein profiles and distinguish between patients with stable and unstable coronary artery disease.

Methods: Patients with ST-segment-elevation myocardial infarction and stable angina pectoris were subjected to routine percutaneous coronary interventions.

The heart arrhythmia-linked D130G calmodulin mutation causes premature inhibitory autophosphorylation of CaMKII.

The Ca/calmodulin (CaM)-dependent kinase II (CaMKII) is well known for transmitting Ca-signals, which leads to a multitude of physiological responses. Its functionality is believed to involve CaMKII holoenzyme dynamics where trans-autophosphorylation of the crucial phosphorylation site, T286 occurs. Phosphorylation of this site does not occur when stimulated exclusively with the arrhythmia associated D130G mutant form of CaM in vitro.

The impact of calmodulin on the cell cycle analyzed in a novel human cellular genetic system.

Calmodulin (CaM) is the principle mediator of the Ca signal in all eukaryotic cells. A huge variety of basic cellular processes including cell cycle control, proliferation, secretion and motility, among many others are governed by CaM, which regulates activities of myriads of target proteins. Mammalian CaM is encoded by three genes localized on different chromosomes all producing an identical protein.

Identification and quantification of sites of nitration and oxidation in the key matrix protein laminin and the structural consequences of these modifications.

Laminin is a major protein of the basement membrane (BM), a specialized extracellular matrix (ECM) of the artery wall. The potent oxidizing and nitrating agent peroxynitrous acid (ONOOH) is formed at sites of inflammation, and data implicate ONOOH in ECM damage and cardiovascular disease. Co-localization of 3-nitrotyrosine, a product of ONOOH-mediated tyrosine (Tyr) modification, and laminin has been reported in human atherosclerotic lesions.

Reaction of quinones with proteins: Kinetics of adduct formation, effects on enzymatic activity and protein structure, and potential reversibility of modifications.

Quinones are a common motif in many biological compounds, and have been linked to tissue damage as they can undergo redox cycling to generate radicals, and/or act as Michael acceptors with nucleophiles, such as protein Cys residues, with consequent adduct formation. The kinetics and consequences of these Michael reactions are poorly characterized. In this study we hypothesized that adduction of protein Cys residues with quinones would be rapid, structure-dependent, quantitatively-significant, and result in altered protein structure and function.

Aggregation of α- and β- caseins induced by peroxyl radicals involves secondary reactions of carbonyl compounds as well as di-tyrosine and di-tryptophan formation.

The present work examined the role of Tyr and Trp in oxidative modifications of caseins, the most abundant milk proteins, induced by peroxyl radicals (ROO). We hypothesized that the selectivity of ROO and the high flexibility of caseins (implying a high exposure of Tyr and Trp residues) would favor radical-radical reactions, and di-tyrosine (di-Tyr) and di-tryptophan (di-Trp) formation. Solutions of α- and β-caseins were exposed to ROO from thermolysis and photolysis of AAPH (2,2'-azobis(2-methylpropionamidine)dihydrochloride).

The peroxyl radical-induced oxidation of Escherichia coli FtsZ and its single tryptophan mutant (Y222W) modifies specific side-chains, generates protein cross-links and affects biological function.

FtsZ (filamenting temperature-sensitive mutant Z) is a key protein in bacteria cell division. The wild-type Escherichia coli FtsZ sequence (FtsZwt) contains three tyrosine (Tyr, Y) and sixteen methionine (Met, M) residues. The Tyr at position 222 is a key residue for FtsZ polymerization.

Investigations of the Navβ1b sodium channel subunit in human ventricle; functional characterization of the H162P Brugada syndrome mutant.

Brugada syndrome (BrS) is a rare inherited disease that can give rise to ventricular arrhythmia and ultimately sudden cardiac death. Numerous loss-of-function mutations in the cardiac sodium channel Nav1.5 have been associated with BrS.