Endo-fucoidan hydrolases from glycoside hydrolase family 107 (GH107) display structural and mechanistic similarities to α-l-fucosidases from GH29

Abstract

Fucoidans are chemically complex and highly heterogeneous sulfated marine fucans from brown macro algae. Possessing a variety of physicochemical and biological activities, fucoidans are used as gelling and thickening agents in the food industry and have anticoagulant, antiviral, antitumor, antibacterial, and immune activities. Although fucoidan-depolymerizing enzymes have been identified, the molecular basis of their activity on these chemically complex polysaccharides remains largely uninvestigated. In this study, we focused on three glycoside hydrolase family 107 (GH107) enzymes: MfFcnA and two newly identified members, P5AFcnA and P19DFcnA, from a bacterial species of the genus Psychromonas. Using carbohydrate-PAGE, we show that P5AFcnA and P19DFcnA are active on fucoidans that differ from those depolymerized by MfFcnA, revealing differential substrate specificity within the GH107 family. Using a combination of X-ray crystallography and NMR analyses, we further show that GH107 family enzymes share features of their structures and catalytic mechanisms with GH29 ␣-L-fucosidases. However, we found that GH107 enzymes have the distinction of utilizing a histidine side chain as the proposed acid/base catalyst in its retaining mechanism. Further interpretation of the structural data indicated that the active-site architectures within this family are highly variable, likely reflecting the specificity of GH107 enzymes for different fucoidan substructures. Together, these findings begin to illuminate the molecular details underpinning the biological processing of fucoidans.

Discovery and Characterization of an Endo-1,3-Fucanase From Marine Bacterium Wenyingzhuangia fucanilytica: A Novel Glycoside Hydrolase Family

“…FunA was incompetent to hydrolyze sulfated fucans from F. vesiculosus , A. nodosum and M. pyrifera ( Supplementary Table S2 ), which were the effective substrates of GH107 enzymes. Currently, the GH107 family is the only reported fucanase GH family, and its enzymes all exhibited endo-1,4-fucanase activity: MfFcnA was specifically active on α-1,4 glycoside bonds in type II sulfated fucans from F. vesiculosus and A. nodosum (the primary structures were shown in Supplementary Figure S2 ; Schultz-Johansen et al, 2018 ); Fp277 and Fp279 could cleave α-1,4 glycoside bonds in A. nodosum ( Schultz-Johansen et al, 2018 ); P5AFcnA and P19DFcnA showed activity on sulfated fucan from M. pyrifera ( Vickers et al, 2018 ), though the primary structure of M. pyrifera was poorly defined. It has not been reported yet whether the current GH107 enzymes could act on α-1,3 glycosidic linkages.…”

“…Schultz-Johansen et al, 2018); Fp277 and Fp279 could cleave α-1,4 glycoside bonds in A. nodosum (Schultz-Johansen et al, 2018); P5AFcnA and P19DFcnA showed activity on sulfated fucan from M. pyrifera (Vickers et al, 2018), though the primary structure of M. pyrifera was poorly defined. It has not been reported yet whether the current GH107 enzymes could act on α-1,3 glycosidic linkages.…”

“…Fucanases are desirable tools to identify the structure of sulfated fucans ( Berteau and Mulloy, 2003 ) and establish structure-activity relationships ( Kusaykin et al, 2016 ). To date, a few genes of endo-1,4-fucanases (i.e., enzymes cleaving α-1,4 glycosidic linkages in type II sulfated fucan backbones) have been characterized, including FcnA, FFA1, FFA2, Fp273, Fp277, Fp279, P5AFcnA, and P19DFcnA ( Colin et al, 2006 ; Silchenko et al, 2017 , 2018 ; Schultz-Johansen et al, 2018 ; Vickers et al, 2018 ). All of the reported endo-1,4-fucanases belong to the glycoside hydrolase (GH) 107 family in the CAZy database 1 .…”

Marine Proteobacteria as a source of natural products: advances in molecular tools and strategies

“…105 Recently, fucoidanases from marine Proteobacteria that are able to degrade brown algae have gained increasingly attention due to the beneficial biomedical properties of fucoidan derived oligosaccharides. 106,107 Heterologous expression has been a powerful tool to identify novel gene functions, but it becomes more cumbersome when gene fragments exceed 10 kb. 25 However, most biosynthetic pathways involve dozens of proteins that catalyze and regulate the production of secondary metabolites and thus are often larger than 10 kb and some may even exceed 100 kb.…”

Enzymatic synthesis of l-fucose from l-fuculose using a fucose isomerase from Raoultella sp. and the biochemical and structural analyses of the enzyme

“…KDH14 isolated from abalone intestine is a novel species that possesses a gene cluster encoding putative l -fucose transporter (FucT), l -fucose mutarotase (FucU), l -fucose isomerase (FucI), l -fuculokinase (FucK), and l -fucose operon activator (FucR), indicating its potential involvement in l -fucose metabolism. Abalone feeds on brown seaweeds containing fucoidan and is a good source of fucoidan-degrading enzymes, which can degrade the polymeric fucoidan into its monomeric l -fucose [25–27]. In this study, Raoultella sp.…”