Interaction between flavin mononucleotide-containing azoreductase and azo dyes. Issue 10 (25th November 2016)

• Record Type: Journal Article
• Authors: Feng, Juan; Wu, Qingqin; Zhou, Yu; Hu, Qizhou; Fang, Ruiqin; Tang, Lixia
• Is Part Of: Spectroscopy letters. Volume 49:Issue 10(2016)
• Page Start: 626
• Page End: 634
• Publication Date: 2016-11-25
• Subjects: Azoreductase -- flavin mononucleotide -- interaction -- molecular docking -- protein engineering; Spectrum analysis -- Periodicals; 543.5
• DOI: 10.1080/00387010.2016.1236822
• Languages: English
• ISSNs: 0038-7010

Abstract

ABSTRACT: Azoreductase, a flavin mononucleotide-containing oxidoreductase from Escherichia coli, can catalyze the reduction of azo dyes to form aromatic amine compounds. Few spectroscopic studies have explored the binding mode of azo dyes or the role of the arginine at site 59 in Azoreductase. In this article, protein engineering strategy has been used to construct one mutant in which the arginine residue at site 59 was mutated to glycine. Fluorescence spectroscopic data showed that the addition of Methyl Red and Methyl Orange resulted in the fluorescence quenching of the cofactor flavin mononucleotide bound to Azoreductase. The association constant was fitted using the standard binding equation instead of the Stern-Volmer equation. The results showed that the mutation from the arginine to glycine at site 59 weakened the association constant from 2.21 × 10 5 L.mol −1 to 4.55 × 10 4 L.mol −1 at 25°C. A similar phenomenon was also observed when Methyl Orange was used as a substrate. In each case, the association constant tended to decrease as the temperature increased from 25°C to 37°C. Thermodynamic parameter analysis revealed that the interaction type changed from a van der Waals interaction (between Azoreductase and the dyes) to a hydrogen bonding interaction (between the mutant and the dyes). Moleculcar docking was also performed in this work to provide some support for the binding mode and binding stability between Azoreductase/mutant and azo dyes.