Laboratory evolution of fungal laccases for the synthesis of hetero-polymeric dyes

Trabajo presentado en el 3rd Multistep Enzyme Catalyzed Processes Congress, celebrado en Madrid (España) del 07 al 10 de abril de 2014.
Fungal laccases (EC1.10.3.2) are blue-multicopper containing enzymes capable of oxidizing phenols, polyphenols, aromatic amines and many other compounds with the concomitant reduction of molecular oxygen to water. The versatility of these enzymes can be expanded by the use of redox mediators (from natural or synthetic origin) thereby finding plenty of applications ranging from bioremediation to novel green processes. Besides, fungal laccases are capable to promote complex hetero-polymerizations of great significance for the synthesis of dyes [1]. The present work describes the engineering of an efficient fungal laccase for the synthesis of polymeric dyes. The departure point for this study was the IG88 mutant from the laccase of Myceliophthora thermophila, which was the genetic product of 21 rounds of directed evolution for secretion in yeast, organic co-solvent tolerance and alkalophilicity [2-4]. With the help of a HTS-assay based on the oxidative cross-coupling (heterocoupling NC) of 2,5-diaminobenzenosulfonic acid and policatechol, laccase mutant libraries were screened and new variants with improved turnover rates under operational conditions were identified