The Discovery of Endo-Fucanases in the GH141 Family: A Novel Functional Activity Within the Family

Brown algae produce structurally complex sulfated fucose-containing polysaccharides known as fucoidans. These compounds are slowly degraded by marine microorganisms, leading to their accumulation in marine sediments and contributing to long-term carbon sequestration. The enzymatic mechanisms underlying fucoidan degradation remain poorly understood. GH141 family enzymes are widely distributed among fucoidan-degrading bacteria, but their function remains hypothetical. It is assumed that during fucoidans degradation, they may act as α-L-fucosidases. We performed a biochemical and bioinformatic analysis of four recombinant enzymes, Wf141_1, Wf141_2, Wf141_3, and Wf141_4, of the GH141 family from the fucoidan-degrading cluster of the marine bacterium <i>Wenyingzhuangia fucanilytica</i> CZ1127^T. Sequence similarity network (SSN) and Conserved Unique Peptide Pattern (CUPP) analysis of the GH141 members revealed that the Wf141s enzymes are distant from previously characterized GH141 members and belong to separate SSN clusters and CUPP branches. All four enzymes exhibited endo-fucanase activity against (1→3;1→4)-α-L-fucoidans. Wf141_1 and Wf141_2 were characterized as sulfated (1→3;1→4)-α-L-fucan endo-1→4-α-L-fucanases (EC 3.2.1.212) with distinct substrate preferences: Wf141_1 preferred [→3-α-L-Fucp2S-1→4-α-L-Fucp2S-1→]_n fragments, whereas Wf141_2 favored [→3-α-L-Fucp2S-1→4-α-L-Fucp2,3S-1→]_n regions. Their specificity depends on structural differences in sugar-binding subsites that recognize sulfation patterns. These enzymes were classified as endo-1→4-α-L-fucanases (EC 3.2.1.212). These findings establish a previously uncharacterized fucoidan-degrading enzymatic function within the GH141 family.