Characterization of a new α-L-fucosidase isolated from the marine mollusk Pecten maximus that catalyzes the hydrolysis of α-L-fucose from algal fucoidan (Ascophyllum nodosum)

Abstract

Algal fucoidan is an α-L-fucose-based polysaccharide endowed with important biological properties for which the structure has not yet been fully elucidated. In an attempt to implement new enzymatic tools for structural study of this polysaccharide, we have found a fucosidase activity in the digestive glands of the common marine mollusk Pecten maximus, which is active on a fucoidan extracted from the brown algae Ascophyllum nodosum. We now report the purification and characterization of this α-L-fucosidase (EC 3.2.1.51). The enzyme was purified by three chromatographic steps, including an essential affinity chromatography based on the glycosidase inhibitor analog 6-amino-deoxy-mannojirimycin as the ligand. The purified α-L-fucosidase is a tetrameric glycoprotein of 200 kDa that hydrolyzes the synthetic substrate p-nitrophenyl α-L-fucopyranoside with a Km value of 650 μM. This enzyme has high catalytic activity (85 μmol · min-1 · mg-1) compared with the other known fucosidases and also possesses an unusual thermal stability. The purified α-L-fucosidase is a retaining glycosidase. The activity of the purified fucosidase was determined on two structurally different fucoidans of the brown algae A. nodosum and Fucus vesiculosus to delineate glycosidic bond specificity. This report is to our knowledge the first demonstration of a fucosidase that can efficiently release α-L-fucose from fucoidan.