The Family Keeps on Growing: Four Novel Fungal OYEs Characterized.

Marina Simona Robescu,Giovanni Loprete,Filippo Vascon,Francesco Filippini,Laura Cendron,Matteo Gasparotto,Elisabetta Bergantino

doi:10.3390/ijms23063050

Marina Simona Robescu, Giovanni Loprete + Show 5 more

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https://doi.org/10.3390/ijms23063050

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Abstract

Aiming at expanding the portfolio of Old Yellow Enzymes (OYEs), which have been systematically studied to be employed in the chemical and pharmaceutical industries as useful biocatalysts, we decided to explore the immense reservoir of filamentous fungi. We drew from the genome of the two Ascomycetes Aspergillus niger and Botryotinia fuckeliana four new members of the OYE superfamily belonging to the classical and thermophilic-like subfamilies. The two BfOYEs show wider substrate spectra than the AnOYE homologues, which appear as more specialized biocatalysts. According to their mesophilic origins, the new enzymes neither show high thermostability nor extreme pH optimums. The crystal structures of BfOYE4 and AnOYE8 have been determined, revealing the conserved features of the thermophilic-like subclass as well as unique properties, such as a peculiar N-terminal loop involved in dimer surface interactions. For the classical representatives BfOYE1 and AnOYE2, model structures were built and analyzed, showing surprisingly wide open access to the active site cavities due to a shorter β6-loop and a disordered capping subdomain.

Highlights

Old Yellow Enzymes (OYEs, EC 1.6.99.1) are a class of flavin-dependent ene-reductases (ERs) catalyzing the asymmetric hydrogenation of electronically activated C=C bonds in the presence of nicotinamide cofactors
Aiming at expanding the portfolio of Old Yellow Enzymes (OYEs), which have been systematically studied to be employed in the chemical and pharmaceutical industries as useful biocatalysts, we decided to explore the immense reservoir of filamentous fungi
We have identified and produced four new putative ERs from the filamentous fungi Aspergillus niger CBS 513.88 and Botryotinia fuckeliana B05.10

Summary

Introduction

Old Yellow Enzymes (OYEs, EC 1.6.99.1) are a class of flavin-dependent ene-reductases (ERs) catalyzing the asymmetric hydrogenation of electronically activated C=C bonds in the presence of nicotinamide cofactors As such, they are biocatalysts sought for effective synthesis methods designed and exploited in the pharmaceutical industry. In 2014, 60 genomes from the clades of Ascomycota and Basidiomycota were widely screened by Nizam et al for the systematic research of OYE homologues, in which 424 putative OYE proteins were identified, and intriguingly, the number of OYE homologues found in each genome varied from 1 to 22 [2] Such an evolutionary diversification was interpreted as a mechanism of fungal adaptation to different environments and carbon sources. The different isoenzymes may have evolved toward the recognition and reduction of specific substrates, as their expression is generally triggered by the surrounding environment (e.g., nutrients, presence of noxious substrates, or chemical-physical parameters) [2,3]

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