Enhanced archaeal laccase production in recombinant Escherichia coli by modification of N-terminal propeptide and twin arginine translocation motifs

摘要: Laccases are multicopper oxidases that couple the oxidation of phenolic polymers to reduction molecular oxygen. While an archaeal laccase has only recently been described (LccA from culture broth Haloferax volcanii), this enzyme appears promising for biotechnology applications based on its robust bilirubin oxidase and activities as well ability withstand prolonged exposure extreme conditions. To further optimize LccA productivity develop option purification whole cells, encoding gene was modified through deletion twin-arginine translocation motif N-terminal propeptide, genes were expressed in Escherichia coli. With approach, readily purified (overall yield up 54 %) soluble fraction E. coli a 74-kDa monomer with syringaldazine oxidizing activity high 33 U mg−1. proteins prepared H. volcanii cells compared by ICP-AES, EPR, DSC, CD, UV–Vis spectroscopy found have similar folding pattern Tm values rich β-sheet structure analogous other oxidases. However, contrast volcanii-purified LccA, which loaded copper, copper not fully incorporated into type-I Cu center thus, providing insight avenues optimization.