Specific detection of quinoproteins by redox-cycling staining.

Paz, Mercedes A.; Flückiger, R; Boak, Andra M.; Kagan, Herbert M.; Gallop, Paul M.

doi:10.1016/s0021-9258(17)35225-0

Cited by 326 publications

(124 citation statements)

References 24 publications

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“…To substantiate the dopaquinone formation in proteins, a redox staining assay was executed using NBT, for which tyrosinase treated samples were transferred onto PVDF membrane after SDS-PAGE. As a result, purple color owing to the redox cycling of dopaquinone and polymerization (shearing of proteins) was observed on the membrane, which corroborates the conversion of DOPA incorporated protein into quinoprotein by the action of tyrosinase [39] (Fig. 3B.4).…”

Section: Coupling Mechanism In Protein For Cell Labelingsupporting

confidence: 67%

Genetically Encoded Dihydroxyphenylalanine Coupled with Tyrosinase for Strain Promoted Labelling

George

Indhu

Sundarapandian

et al. 2021

Preprint

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Protein modifications through the genetic code engineering method have a remarkable impact on macromolecule engineering, protein translocation, protein-protein interaction, and cell biology. Here, the newly developed molecular biology approach expanding the genetic code was used for fine tuning of protein for biological availability. For that non-canonical amino acid dihydroxyphenylalanine was genetically incorporated at the defined site in the protein. Further, the congener protein was enzymatically controlled for direct conversion of quinone for strain promoted click chemistry reaction. It yields a single product with defined stereochemistry and temporally controlled conjugation with BCN. The feasibility was explored for selective cell imaging and programmed cell death in HeLa cells.

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Section: Coupling Mechanism In Protein For Cell Labelingsupporting

confidence: 67%

Genetically Encoded Dihydroxyphenylalanine Coupled with Tyrosinase for Strain Promoted Labelling

George

Indhu

Sundarapandian

et al. 2021

Preprint

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“…It is based on the ability of DOPA-containing proteins to reduce nitroblue tetrazolium in the presence of alkaline pH and an excess of glycine (used as reducing agent). The resulting product, formazan, is a deep blue coloured compound [134]. Both methods were used to visualize the distribution of DOPA-containing proteins in the cement cells of the tubeworms P. californica [34,56] and S. alveolata [36].…”

Section: Hydroxylationmentioning

confidence: 99%

Experimental strategies for the identification and characterization of adhesive proteins in animals: a review

et al. 2015

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Adhesive secretions occur in both aquatic and terrestrial animals, in which they perform diverse functions. Biological adhesives can therefore be remarkably complex and involve a large range of components with different functions and interactions. However, being mainly protein based, biological adhesives can be characterized by classical molecular methods. This review compiles experimental strategies that were successfully used to identify, characterize and obtain the full-length sequence of adhesive proteins from nine biological models: echinoderms, barnacles, tubeworms, mussels, sticklebacks, slugs, velvet worms, spiders and ticks. A brief description and practical examples are given for a variety of tools used to study adhesive molecules at different levels from genes to secreted proteins. In most studies, proteins, extracted from secreted materials or from adhesive organs, are analysed for the presence of post-translational modifications and submitted to peptide sequencing. The peptide sequences are then used directly for a BLAST search in genomic or transcriptomic databases, or to design degenerate primers to perform RT-PCR, both allowing the recovery of the sequence of the cDNA coding for the investigated protein. These sequences can then be used for functional validation and recombinant production. In recent years, the dual proteomic and transcriptomic approach has emerged as the best way leading to the identification of novel adhesive proteins and retrieval of their complete sequences.

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“…present in myocardium. Therefore, we refined a technique using nitroblue tetrazolium (NBT), to determine whether myocardial tissue contains catechol-modified protein adducts (51). NBT is a dye that selectively reacts with the catechol moiety on proteins to form a stable blue conjugate on nitrocellulose membrane (Fig.…”

Section: Innovationmentioning

confidence: 99%

“…NBT is a redox-cycling dye that binds to catechol-protein adducts within a nitrocellulose membrane and stains the proteins blue, allowing the visualization of catechol-modified proteins (51,52). Myocardial lysate alone, or after having been incubated with catecholamines -MAO inhibitors, was subjected to SDS-PAGE.…”

Section: Detection and Proteomic Analysis Of Catecholaldehydemodified Proteinsmentioning

confidence: 99%

Enhanced Catecholamine Flux and Impaired Carbonyl Metabolism Disrupt Cardiac Mitochondrial Oxidative Phosphorylation in Diabetes Patients

Nelson

Efird

Kew

et al. 2021

Antioxidants & Redox Signaling

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Aims: Catecholamine metabolism via monoamine oxidase (MAO) contributes to cardiac injury in models of ischemia and diabetes, but the pathogenic mechanisms involved are unclear. MAO deaminates norepinephrine (NE) and dopamine to produce H 2 O 2 and highly reactive ''catecholaldehydes,'' which may be toxic to mitochondria due to the localization of MAO to the outer mitochondrial membrane. We performed a comprehensive analysis of catecholamine metabolism and its impact on mitochondrial energetics in atrial myocardium obtained from patients with and without type 2 diabetes. Results: Content and maximal activity of MAO-A and MAO-B were higher in the myocardium of patients with diabetes and they were associated with body mass index. Metabolomic analysis of atrial tissue from these patients showed decreased catecholamine levels in the myocardium, supporting an increased flux through MAOs. Catecholaldehyde-modified protein adducts were more abundant in myocardial tissue extracts from patients with diabetes and were confirmed to be MAO dependent. NE treatment suppressed mitochondrial ATP production in permeabilized myofibers from patients with diabetes in an MAO-dependent manner. Aldehyde dehydrogenase (ALDH) activity was substantially decreased in atrial myocardium from these patients, and metabolomics confirmed lower levels of ALDH-catalyzed catecholamine metabolites. Proteomic analysis of catechol-modified proteins in isolated cardiac mitochondria from these patients identified >300 mitochondrial proteins to be potential targets of these unique carbonyls. Innovation and Conclusion: These findings illustrate a unique form of carbonyl toxicity driven by MAOmediated metabolism of catecholamines, and they reveal pathogenic factors underlying cardiometabolic disease. Importantly, they suggest that pharmacotherapies targeting aldehyde stress and catecholamine metabolism in heart may be beneficial in patients with diabetes and cardiac disease. Antioxid. Redox Signal. 35,[235][236][237][238][239][240][241][242][243][244][245][246][247][248][249][250][251]

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Specific detection of quinoproteins by redox-cycling staining.

Cited by 326 publications

References 24 publications

Genetically Encoded Dihydroxyphenylalanine Coupled with Tyrosinase for Strain Promoted Labelling

Genetically Encoded Dihydroxyphenylalanine Coupled with Tyrosinase for Strain Promoted Labelling

Experimental strategies for the identification and characterization of adhesive proteins in animals: a review

Enhanced Catecholamine Flux and Impaired Carbonyl Metabolism Disrupt Cardiac Mitochondrial Oxidative Phosphorylation in Diabetes Patients

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