Lignin-degrading peroxidases in white-rot fungus <i>Trametes hirsuta</i> 072. Absolute expression quantification of full multigene family

<div><p>Ligninolytic heme peroxidases comprise an extensive family of enzymes, which production is characteristic for white-rot Basidiomycota. The majority of fungal heme peroxidases are encoded by multigene families that differentially express closely related proteins. Currently, there were very few attempts to characterize the complete multigene family of heme peroxidases in a single fungus. Here we are focusing on identification and characterization of peroxidase genes, which are transcribed and secreted by basidiomycete <i>Trametes hirsuta</i> 072, an efficient lignin degrader. The <i>T</i>. <i>hirsuta</i> genome contains 18 ligninolytic peroxidase genes encoding 9 putative lignin peroxidases (LiP), 7 putative short manganese peroxidases (MnP) and 2 putative versatile peroxidases (VP). Using ddPCR method we have quantified the absolute expression of the 18 peroxidase genes under different culture conditions and on different growth stages of basidiomycete. It was shown that only two genes (one MnP and one VP) were prevalently expressed as well as secreted into cultural broth under all conditions investigated. However their transcriptome and protein profiles differed in time depending on the effector used. The expression of other peroxidase genes revealed a significant variability, so one can propose the specific roles of these enzymes in fungal development and lifestyle.</p></div

The <i>de novo</i> sequenced peptides of JnL and SbL aligned with the amino acid sequences of laccase 2 from <i>S</i>. <i>murashkinskyi</i> (SmL) and laccase A from <i>A</i>. <i>faginea</i> (AfL).

<p>The glycosylated peptides are shown in red. The potential glycosylation sites are highlighted in grey, and the occupied glycosylation sites are highlighted in red. The coordinating ligands of the copper ions are highlighted in turquoise.</p

Oxidative potential of <i>Junghuhnia nitida</i> and <i>Steccherinum bourdotii</i> strains evaluated by the express method.

<p>Oxidative potential of <i>Junghuhnia nitida</i> and <i>Steccherinum bourdotii</i> strains evaluated by the express method.</p

The kinetic parameters of oxidation of the various substrates by the laccases.

<p>The kinetic parameters of oxidation of the various substrates by the laccases.</p

The activity profiles of the lignin degrading enzymes during <i>J</i>. <i>nitida</i> (red circles) and <i>S</i>. <i>bourdotii</i> (black squares) static surface cultivation on LC medium.

<p>Panels A-C show the activities of laccase, manganese peroxidase and Mn²⁺- independent peroxidases (versatile peroxidase and/or lignin peroxidase), respectively.</p

Physicochemical characteristics of the laccases.

<p>Physicochemical characteristics of the laccases.</p