Official Full Name
Cyclohexanone Monooxygenase
Background
Purified cyclohexanone monooxygenase is a versatile oxygenation catalyst. The enzyme uses the bound FAD-4a-OOH oxygenating intermediate to initiate transfer of oxygen to electrophilic substrate sites. The reaction consequently yields the corresponding sulfoxide and selenoxide products. This enzyme is also capable of oxygenating at nitrogen, trivalent phosphorus, and boron sites in boronic acids. Hence, it is one of the most broad-based flavoprotein oxygenases known.
Synonyms
yclohexanone 1#2-monooxygenase; cyclohexanone oxygenase; cyclohexanone:NADPH:oxygen oxidoreductase (6-hydroxylating# 1#2-lactonizing); cyclohexanone monooxygenase; EC 1.14.13.22; 52037-90-8; cyclohexanone#NADPH:oxygen oxidoreductase (lactone-forming)
Introductions
Cyclohexanone monooxygenase is a flavin-dependent monooxygenase with an extremely broad substrate spectrum, capable of catalyzing more than 100 different reactions such as the oxygenation of heteroatoms of nitrogen, boron and sulfur. Compared with chemical catalysts, Cyclohexanone monooxygenase can directly use molecular oxygen as oxidant in catalysis, and it can achieve high stereoselectivity under milder conditions, so it is now an indispensable tool enzyme in organic synthesis.
CHMO
Cyclohexanone monooxygenase (CHMO), which is derived from the genus Bacillus, is the first CHMO with potential for synthesis. Cyclohexanone monooxygenase has a molecular mass of about 60 kDa and is capable of catalyzing more than 100 substrates such as cyclic ketones and aldehydes, with a very broad substrate spectrum. Besides, CHMO is also capable of catalyzing heteroatoms, such as sulfides, tertiary amines, sulfites, etc., with very powerful catalytic ability and selectivity.
Classifications
CHMO enzymes are classified into three categories according to their coenzymes dependence. Tpye I CHMO depends on flavin and NADPH as cofactors and belongs to group B flavin monooxygenases. Tpye II CHMO depends on NADH and FAD and belongs to class C flavin monooxygenases. Tpye III CHMO is atypical CHMO belongs to class A flavin monooxygenases. Among them, CHMO belongs to class I CHMO, which depends on flavin adenine dinucleotide (FAD) and NADPH as coenzymes. This class of CHMO contains two characteristic fingerprint sequences, "GXGXXG", which are conserved in two structural domains and are non-covalent binding sites for the coenzymes FAD and NADPH, respectively. CHMO contains two structural domains, FAD-binding domain and NADPH-binding domain. Mutation of amino acids in this binding domain can change its coenzyme dependence. mutation of important residues for NADPH binding by Fraaije et al. changed the mutant from NADPH-dependent to NADH-dependent.
Catalytic mechanism
Throughout the enzymatic reaction catalyzed by CHMO, the coenzyme NADPH first reduces FAD to form the reduced state of FAD, which is oxidized by molecular oxygen to form the C4α-peroxyisocyanine intermediate. This intermediate acts as a nucleophilic reagent to attack the carbonyl carbon atom on the substrate bound to the substrate pocket, forming a tetrahedral "Criegee" intermediate, followed by molecular rearrangement, transferring the oxygen atom to the substrate in the Baeyer-Villiger reaction. After the rearrangement, a molecule of water is shed from the C4α-peroxyisocyanine intermediate and the FAD is regenerated, and NADP is bound to the C4α-peroxyisocyanine intermediate throughout the reaction to maintain the stability of the intermediate.
Biological Function
CHMO is a bacterial flavoenzyme whose main function in the cell is to catalyze the conversion of cyclohexanone, a cyclic ketone, into ε-caprolactone, a key step in the pathway for the biodedgredation of cyclohexanol. However, given the lack of specificity for CHMO, it can be used generally to form lactones from a number of four to six-membered cyclic ketones, which can then be hydrolyzed into aliphatic acids. Moreover, CHMO has the ability to oxygenate aromatic aldehydes and heteroatom-containing compounds – such as trivalent phosphorus and boronic acids– as well, making it a candidate for industrial use.
Enzyme Mechanism
Cyclohexanone monooxygenase (CHMO) uses NADPH and O2 as cosubstrates and FAD as a cofactor to insert an oxygen atom into the substrate. The process involves the formation of a falvin-peroxide and Criegee intermediate.
