Purification and characterization of a fungal laccase expressed in Kluyveromyces lactis suitable for baking.

ABSTRACT

Laccase enzyme can replace chemical additives to improve texture properties and the volume of bread. Laccase encoding gene from Phlebia brevispora, a native fungus from Misiones, Argentina, was expressed in the generally recognized as safe yeast Kluyveromyces lactis. To improve laccase activity, medium conditions were optimized. The use of iron sulfate at a concentration of 1 mM led to optimum laccase activity (1289 U·L-1 ) on the fourth day of incubation. SDS-PAGE analysis revealed that the molecular mass of purified laccase was about 180 kDa. Optimum pH for the enzyme was 4 and optimum temperature was 40°C. Laccase exhibited high stability at low pH and high temperature. The application of recombinant laccase to bread decreased hardness, gumminess, and chewiness and increased bread volume. Based on these results, recombinant laccase from P. brevispora with improved yield is a good option for application as an improver of the physicochemical quality of bread at the industrial level. Besides, it will allow us to advance toward our goal of developing healthy alternatives for the bakery industry. No previous work has been reported concerning the heterologous expression of the laccase gene native to the province of Misiones, Argentina, with an aim for application in baking. PRACTICAL APPLICATION Healthy bakeries became a trend in recent years. The use of the laccase enzyme increases the specific volume and decreases the hardness of bread, being thus an alternative for the replacement of chemical additives in the bakery industry.