Tailoring a Soluble Diiron Monooxygenase for Synthesis of Aromatic N-oxides

Narimantas Čėnas,Rolandas Meškys,Vytautas Petkevičius,Justas Vaitekūnas,Dovydas Vaitkus

doi:10.3390/catal9040356

Abstract

The aromatic N-oxides have received increased attention over the last few years due to their potential application in medicine, agriculture and organic chemistry. As a green alternative in their synthesis, the biocatalytic method employing whole cells of Escherichia coli bearing phenol monooxygenase like protein PmlABCDEF (from here on – PML monooxygenase) has been introduced. In this work, site-directed mutagenesis was used to study the contributions of active site neighboring residues I106, A113, G109, F181, F200, F209 to the regiospecificity of N-oxidation. Based on chromogenic indole oxidation screening, a collection of PML mutants with altered catalytic properties was created. Among the tested mutants, the A113G variant acquired the most distinguishable N-oxidations capacity. This new variant of PML was able to produce dioxides (quinoxaline-1,4-dioxide, 2,5-dimethylpyrazine-1,4-dioxide) and specific mono-N-oxides (2,3,5-trimethylpyrazine-1-oxide) that were unachievable using the wild type PML. This mutant also featured reshaped regioselectivity as N-oxidation shifted towards quinazoline-1-oxide compared to quinazoline-3-oxide that is produced by the wild type PML.

Highlights

Biocatalysis has become an attractive alternative to chemical synthesis because of its exceptional selectivity, high efficiency and ability to produce relatively pure compounds
Most recently we have demonstrated that an enzyme of SDIMO family could be employed to catalyze a different type of oxidation as phenol monooxygenase like (PML) monooxygenase transformed numerous nitrogen-containing aromatic compounds to the corresponding N-oxides [11]
The wild type PML has been introduced as a novel biocatalyst for oxidation of

Summary

Introduction

Biocatalysis has become an attractive alternative to chemical synthesis because of its exceptional selectivity, high efficiency and ability to produce relatively pure compounds. Oxygenases catalyze a wide variety of reactions including activation of sp hybridized C atoms, epoxidation of C=C double bonds, aromatic hydroxylation, N-oxidation, deamination and dehalogenation, Baeyer-Villiger oxidation, as well as N-, O- and S-dealkylation [5]. These enzymes can accept a diversity of substrates, including fatty acids, terpenes, steroids, prostaglandins, mono-aromatic, poly-aromatic and heteroaromatic compounds, as well as alkanes, alkenes, organic solvents, antibiotics, pesticides, carcinogens and Catalysts 2019, 9, 356; doi:10.3390/catal9040356 www.mdpi.com/journal/catalysts

Methods

Results

Conclusion