(48e) Site-Specific Cross-Linking and Immobilization of Agarase Enzymes for Conversion of Gelidium Amansii into Biologically-Active Sugars

Cuevas, R. A. I. - Presenter, Myongji University
Weldemhret, T. G. K., Myongji University
Valdehuesa, K. N. G., Myongji University
Nisola, G., Myongji University
Fissaha, H. T., Myongji University
Lee, W. K., Myongji University
Chung, W. J., Myongji University
Ramos, K. R. M., Myongji University
Red macroalgae Gelidium amansii is currently considered as a prospective renewable resource due to its ubiquity, high carbohydrate and low lignin contents. Its major cell wall carbohydrate is agarose, and sugars derived from agarose have various industrial potentials, i.e. D-galactose can be fermented to ethanol whereas neoagaro-oligosaccharides, neoagarobiose, and 3,6-anhydro-L-galactose can be utilized as biofunctional materials owing to their anticariogenic, anti-inflammation, probiotic, skin whitening and moisturizing properties. Selective production of these sugars without degradation is needed to preserve their bio-functionality. In this context, G. amansii can be effectively pretreated by using ionic liquids (ILs) as non-derivatizing solvents combined with enzymatic hydrolysis. However, its large scale feasibility is still challenged due to the high cost of enzymes (i.e. agarases), which have low thermostability and IL tolerance. Herein, tyrosinase-catalyzed site-specific cross-linking and covalent immobilization of agarase (i.e. Aga2 and AhgI from C. omnivescoria sp. nov. W5C and Aga50D from S. degradans 2-40) is proposed to tackle these challenges. Tyrosine residues (Y-tag) were introduced to the enzymes via genetic engineering, and mushroom tyrosinase was utilized to catalyze the crosslinking and immobilization reactions. Site-specific crosslinking and immobilization of agarases improve their activities, thermal stabilities, IL tolerance and recyclability for repeated production of agarose-derived sugars. This work was supported by the Basic Science Research Program and Korea Research Fellowship Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Science and ICT (No. 2015H1D3A1062172 and 2016R1C1B1013252), and the Ministry of Education (No. 22A20130012051(BK21Plus)).