Na+/K+-ATPase interaction with methylglyoxal as reactive metabolic side product
Language English Country United States Media print-electronic
Document type Journal Article
PubMed
28342847
DOI
10.1016/j.freeradbiomed.2017.03.024
PII: S0891-5849(17)30173-9
Knihovny.cz E-resources
- Keywords
- Aminoguanidine, Enzyme inhibition, Mass spectrometry, Methylglyoxal, Oxidative post-translational modification, Reactivity, Sodium pump,
- MeSH
- Guanidines pharmacology MeSH
- Mass Spectrometry MeSH
- Protein Conformation MeSH
- Crystallography, X-Ray MeSH
- Kidney metabolism MeSH
- Ouabain pharmacology MeSH
- Oxidative Stress MeSH
- Glycation End Products, Advanced chemistry metabolism MeSH
- Pyruvaldehyde chemistry metabolism MeSH
- Serum Albumin, Bovine metabolism MeSH
- Cattle MeSH
- Sodium-Potassium-Exchanging ATPase antagonists & inhibitors chemistry metabolism MeSH
- Sus scrofa MeSH
- Protein Binding MeSH
- Animals MeSH
- Check Tag
- Cattle MeSH
- Animals MeSH
- Publication type
- Journal Article MeSH
- Names of Substances
- Guanidines MeSH
- Ouabain MeSH
- pimagedine MeSH Browser
- Glycation End Products, Advanced MeSH
- Pyruvaldehyde MeSH
- Serum Albumin, Bovine MeSH
- Sodium-Potassium-Exchanging ATPase MeSH
Proteins are subject to oxidative modification and the formation of adducts with a broad spectrum of reactive species via enzymatic and non-enzymatic mechanisms. Here we report that in vitro non-enzymatic methylglyoxal (MGO) binding causes the inhibition and formation of MGO advanced glycation end-products (MAGEs) in Na+/K+-ATPase (NKA). Concretely, MGO adducts with NKA amino acid residues (mainly Arg) and Nε-(carboxymethyl)lysine (CML) formation were found. MGO is not only an inhibitor for solubilized NKA (IC50=91±16μM), but also for reconstituted NKA in the lipid bilayer environment, which was clearly demonstrated using a DPPC/DPPE liposome model in the presence or absence of the NKA-selective inhibitor ouabain. High-resolution mass spectrometric analysis of a tryptic digest of NKA isolated from pig (Sus scrofa) kidney indicates that the intracellular α-subunit is naturally (post-translationally) modified by MGO in vivo. In contrast to this, the β-subunit could only be modified by MGO artificially, and the transmembrane part of the protein did not undergo MGO binding under the experimental setup used. As with bovine serum albumin, serving as the water-soluble model, we also demonstrated a high binding capacity of MGO to water-poorly soluble NKA using a multi-spectral methodology based on electroanalytical, immunochemical and fluorimetric tools. In addition, a partial suppression of the MGO-mediated inhibitory effect could be observed in the presence of aminoguanidine (pimagedine), a glycation suppressor and MGO-scavenger. All the results here were obtained with the X-ray structure of NKA in the E1 conformation (3WGV) and could be used in the further interpretation of the functionality of this key enzyme in the presence of highly-reactive metabolic side-products, glycation agents and generally under oxidative stress conditions.
References provided by Crossref.org
