Heterologous expression of a Streptomyces cyaneus laccase for biomass modification applications

Selin Ece,Rainer Fischer,Camilla Lambertz,Ulrich Commandeur

doi:10.1186/s13568-017-0387-0

Abstract

Laccases are used for the conversion of biomass into fermentable sugars but it is difficult to produce high yields of active laccases in heterologous expression systems. We overcame this challenge by expressing Streptomyces cyaneus CECT 3335 laccase in Escherichia coli (ScLac) and we achieved a yield of up to 104 mg L−1 following purification by one-step affinity chromatography. Stability and activity assays using simple lignin model substrates showed that the purified enzyme preparation was active over a broad pH range and at high temperatures, suggesting it would be suitable for biomass degradation. The reusability of ScLac was also demonstrated by immobilizing the enzyme on agarose beads with a binding yield of 33%, and by the synthesis of cross-linked enzyme aggregates with an initial activity recovery of 72%.

Highlights

Laccases are the largest subgroup of the multi-copper oxidase protein superfamily (Ihssen et al 2015)
Each monomeric laccase contains four copper atoms located at three different positions, namely the type 1 (T1), type 2 (T2) and binuclear type 3 (T3) copper sites, all of which are involved in the oxidation of substrate molecules accompanied by the reduction of molecular oxygen to two molecules of water (Thurston 1994)
Analysis of the enzymatic activity and functional properties of recombinant ScLac The visible spectra of the purified ScLac correlated with the typical spectra of blue laccases

Summary

Introduction

Laccases are the largest subgroup of the multi-copper oxidase protein superfamily (Ihssen et al 2015). They can oxidize a broad range of substrates including phenolic compounds, azo dyes, aromatic amines, non-phenolic substrates (mostly with the help of mediators), anilines and aromatic thiols, and recalcitrant environmental pollutants (Canas and Camarero 2010; Majumdar et al 2014; Margot et al 2013; Widsten and Kandelbauer 2008). The digestibility of cellulose can be increased following lignin decomposition by laccases (Chen et al 2012). In the presence of HBT, lignin can be removed from whole woody and nonwoody feedstocks to increase sugar and ethanol yields (Gutierrez et al 2012), whereas the alternative mediator

Methods

Results

Conclusion