LAC3, a new low redox potential laccase from Trametes sp. strain C30 obtained as a recombinant protein in yeast

Trametes sp. strain C30 modulates the synthesis of various laccase isoforms in response to external stimuli. Systematic cloning and heterologous expression of laccase cDNAs were carried out to find C30 enzymes with high oxidative capacities. Two cDNAs were successfully expressed in the yeast Saccharomyces cerevisiae:clac1, encoding the previously characterized LAC1 and clac3, encoding a previously undetected laccase isoform. The amino acid sequence deduced from clac3 reveals a protein of 524 amino acids (estimated pI = 4.0) that is, respectively, 67 and 75% identical (79 and 84% similar) to LAC1 and LAC2. Up to 2 mg/L of recombinant LAC3 were produced from a yeast fermentor culture. LAC3 recombinant enzyme properties are very close to those of the inducible native LAC2: it contains a low potential T1 copper (E° = 0.53 V) and is remarkably active both on phenolic (kcat syringaldazine = 56,600 min−1 and kcat guaïacol = 43,250 min−1) and non-phenolic (kcat ABTS = 56,650 min−1) substrates. This work shows that laccases weakly expressed in white-rot fungi can however be studied after a functional expression in yeast.