Peculiarities of hydroxyaryl oxidation by fungal peroxidase: reactivity and inhibition of the enzyme

Abstract

The oxidation kinetics of hydroxyaryls (HAs), i. e. phenol, 1-naphthol, 2-naphthol, 4,4’-isopropylidene diphenol (bisphenol A), 4-hydroxybiphenyl and 1-hydroxypyrene, catalysed with the recombinant Coprinus cinereus peroxidase (rCiP) was investigated with an emphasis on the correlation between the structure and the function of the substrates. The HAs showed different reactivity. The oxidation rate of HAs was maximal at pH 7.0–7.5. For 1-naphthol, an apparent pKa of activity change was 5.6 and 9.4. The apparent bimolecular constant of HA oxidation (kb) changed from 1.3 • 105 to 1.3 • 108 M–1s–1 at pH 5.5 and 25 oC. Comparison of the initial rate of HA oxidation with the calculated molecular parameters has shown that the rate-determining factors are the energy of molecular orbitals (redox potential) and the hydrophobicity of the substrates. During exhaustive HA oxidation, peroxidase inhibition and microparticle formation were indicated. BSA and PEG prevented the inhibition and increased the oxidation yield of HA. A hypothesis was made that inhibition of rCiP is linked with formation of HA oligomer which bind in the active centre of peroxidase. The docking calculations show that these oligomers interact with the enzyme stronger than the substrates. The oligomers do not form productive complexes and block the active centre. BSA, PEG and other non-charged polymers that form a globular structure bind oligomers and prevent enzyme inhibition. Keywords: Coprinus cinereus, peroxidase, phenol, 1-naphthol, 2-naphthol, 4-hydroxybiphenyl, bisphenol A, 1-hydroxypyrene, BSA, PEG