Abstrakt
alpha-L-Rhamnosidase (EC 3.2.1.40) is a biotechnologically important enzyme used for derhamnosylation of many natural compounds. The extracellular alpha-L-rhamnosidase was purified from the culture of Aspergillus terreus grown on L-rhamnose-rich medium.
This enzyme was found to be thermo- and alkali-tolerant, able to operate at 70 degrees C and pH 8.0. The alpha-L-rhamnosidase cDNA was cloned from A. terreus, sequenced, and expressed in the yeast Pichia pastoris as a fully functional protein.
The recombinant protein was purified to apparent homogeneity and biochemically characterized. Both the native and the recombinant alpha-L-rhamnosidases catalyzed the conversion of rutin into quercetin-3-glucopyranoside (isoquercitrin), a pharmacologically significant flavonoid usable in nutraceutics.
This procedure has high volumetric productivity (up to 300 g/L) and yields the product void of unwanted quercetin. The significant advantage of our expression system consists in shorter production times, up to fourfold increase in enzyme yields and the absence of unwanted beta-D-glucosidase as compared to the native production system.
Thanks to its unique properties, this enzyme is applicable in a selective synthesis/hydrolysis of various rhamnose containing structures.