The amplified detection of free and bound methoxatin (PQQ) with nitroblue tetrazolium redox reactions: insights into the PQQ-locus.
Porcine kidney diamine oxidase, a PQQ-enzyme, can be directly measured by formazan production with putrescine and nitroblue tetrazolium. This cyclic reaction in air is unaffected by superoxide dimutase, suggesting a two electron transfer between substrate-reduced PQQ-locus and nitroblue tetrazolium, without intermediate formation of superoxide. With albumin-bound PQQ and detergent-exposed PQQ-loci, glycine can be oxidized by PQQ and electrons repetitively transferred through PQQ-sites to nitroblue tetrazolium, the rate of formazan production detecting picomoles of exposed PQQ-locus. Exposed PQQ-loci are also reducible with NaCNBH3. Nitroblue tetrazolium, reoxidizes the reduced PQQ-locus with formazan production. These experiments suggest that the PQQ-locus of quinoproteins contains a [ketone-ketoimine in equilibrium with ketoamine] redox center.
D006631:Amine Oxidase (Copper-Containing) / Q000378:metabolism*; D000818:Animals; D007202:Indicators and Reagents; D007668:Kidney / Q000201:enzymology; D007700:Kinetics; D008832:Microchemistry; D009580:Nitroblue Tetrazolium; D010084:Oxidation-Reduction; D045542:PQQ Cofactor; D011485:Protein Binding; D011700:Putrescine / Q000378:metabolism; D011804:Quinolines / Q000032:analysis* / Q000378:metabolism; D013053:Spectrophotometry / Q000379:methods; D013552:Swine
D007202:Indicators and Reagents; D011804:Quinolines; D009580:Nitroblue Tetrazolium; D045542:PQQ Cofactor; D006631:Amine Oxidase (Copper-Containing); D011700:Putrescine