Catalytic properties of a short manganese peroxidase from Irpex lacteus F17 and the role of Glu166 in the Mn2+-independent activity

Abstract

Il-MnP1 (GenBank: AGO86670.2) has been confirmed by sequence analysis as a short manganese peroxidase (MnP) from Irpex lacteus F17 (CCTCC AF 2014020). To investigate the catalytic properties, the oxidation of typical aromatic substrates and the pathways of guaiacol oxidation by Il-MnP1, both in the presence and absence of Mn2+ at either pH 4.0 or pH 7.4, were analyzed. Results showed that Il-MnP1 exhibited higher oxidative activity in the presence of Mn2+ than in the absence of Mn2+ toward the majority of the selected substrates at pH 4.0. Additionally, the similar product compositions suggested that the oxidation of guaiacol mainly belongs to a series of polymeric reactions of radicals initiated by Il-MnP1, whether they were in the presence and absence of Mn2+ at either pH 4.0 or 7.4. Furthermore, two variants (E166G, E166Q) were found using site-directed mutagenesis, to improve the Mn2+-independent oxidative activity significantly. The catalytic efficiency (Kcat/Km) of E166G and E166Q in 2, 6-dimethoxyphenol oxidation was higher than Il-MnP1 by 170 and 34 times, respectively. The study revealed certain differences in catalytic properties between Mn2+ dependent and independent oxidation by Il-MnP1. More importantly, a residue (E166) was related to the Mn2+-independent activity of a short MnP.