Enzymatic production of 2'-fucosyllactose from l-fucose derived from acid-hydrolyzed Ishige foliacea using a bifunctional fucokinase/l-fucose 1-l-P guanyltransferase system.

Brown algae have emerged as a promising natural source of fucose for the production of 2'fucosyllactose (2'-FL) and fucoidan. This study presents an efficient enzymatic method for producing 2'-FL from l-fucose derived from Ishige foliacea using a bifunctional enzyme system. Among the 22 brown algae species found in Korea, I. foliacea was identified as the most abundant source of fucose, containing 12.33 ± 0.22 % of fucose. l-Fucose was extracted via optimized acid hydrolysis and purified through electrodialysis, selective fermentation with Saccharomyces cerevisiae, Scheffersomyces stipitis, and activated carbon absorption. The bifunctional enzyme system consisted of recombinant l-fucokinase/l-fucose-1-P guanyltransferase (FKP) from Bacteroides fragilis and α1,2-fucosyltransferase (FucT) from Helicobacter pylori, both expressed in Escherichia coli Rosetta. The FucT enzymes were tagged with trigger factor (TF), which significantly enhanced their solubility and activity. Under optimized conditions (pH 8.5, 27 °C, 55 μM of enzyme), the system achieved a 2'-FL conversion yield of 50.21 ± 1.03 % after 48 h. Mass balance analysis revealed a final 2'-FL production of 13.64 g (27.93 mM) from 9.13 g of fucose and 19.04 g of lactose, demonstrating the practical feasibility of producing 2'-FL from I. foliacea-derived fucose through this bifunctional enzymatic approach.