Role and structure of the small subunit forming heterodimers with laccase‐like enzymes

Abstract

Unlike laccases sensu stricto, which are usually monomeric enzymes, laccase‐like enzymes recently re‐classified as Novel Laccases (NLACs) are characterized by the formation of heterodimers with small proteins (subunits) of unknown function. Here the NLAC from Pleurotus eryngii (PeNL) and a small protein selected from the fungal genome, that is homologous to reported POXA3 from Pleurotus ostreatus, were produced in Aspergillus oryzae separately or together. The two proteins interacted regardless of whether the small subunit was co‐expressed or exogenously added to the enzyme. The stability and catalytic activity of PeNL was significantly enhanced in the presence of the small subunit. SEC‐MALS analysis confirmed that the complex PeNL‐ss is a heterodimer of 77.4 kDa. The crystallographic structure of the small protein expressed in Escherichia coli was solved at 1.6 Å resolution. This is the first structure elucidated of a small subunit of a NLAC. The helix bundle structure of the small subunit accommodates well with the enzyme model structure, including interactions with specific regions of NLACs and some amino acid residues of the substrate‐binding loops.This article is protected by copyright. All rights reserved.

Fungal Laccases: Fundamentals, Engineering and Classification Update

“…Moreover, the crystal structure of a small subunit was solved for the first time. Finally, the observed interactions between the catalytic and the small subunit indicated that the NLAC holds structural features likely involved in substrate binding and/or the interaction with the small subunit, which could explain the differences in activity and stability of monomeric or complexed NLACs, as well as of NLACs compared to laccases sensu stricto [77].…”

“…This has been recently confirmed during the expression and characterization of the heterodimeric complex formed by the NLAC of P. eryngii with a small protein identified in the genome of the fungus. In that study, the stability to pH, temperature and presence of organic co-solvents and the catalytic activity of the enzyme was remarkably increased by the presence of the small subunit [77]. Moreover, the crystal structure of a small subunit was solved for the first time.…”