title: Expression and biochemical characterization and substrate specificity of the fucoidanase from Formosa algae.
abstract: A gene that encodes fucoidanase ffa2 in the marine bacterium Formosa algae strain KMM 3553T was cloned, and the protein (FFA2) was produced in Escherichia coli. Recombinant fucoidanase FFA2 was purified, and the biochemical properties of this enzyme were studied. The amino acid sequence of FFA2 showed 57% identity with known fucoidanase FcnA from Mariniflexile fucanivorans. The mass of the gene product FFA2 is 101.2 kDa (918 amino acid residues). Sequence analysis has revealed that fucoidanase FFA2 belongs to the GH107 (CAZy) family. Detailed substrate specificity was studied by using fucoidans from brown seaweeds as well as synthetic fucooligosaccharide with distinct structures. Fucoidanase FFA2 catalyzes the cleavage of (1→4)-α-glycosidic bonds in the fucoidan from Fucus evanescens within a structural fragment (→3)-α-l-Fucp2S-(1→4)-α-l-Fucp2S-(1→)n but not in a fragment (→3)-α-l-Fucp2S,4S-(1→4)-α-l-Fucp2S-(1→)n. Using synthetic di-, tetra- and octasaccharides built up of the alternative (1→4)- and (1→3)-linked α-l-Fucp2S units, the difference in substrate specificity and in the rate of enzymatic selectivity was investigated. Nonsulfated and persulfated synthetic oligosaccharides were not transformed by the enzyme. Therefore, FFA2 was specified as poly[(1→4)-α-l-fucoside-2-sulfate] glycanohydrolase. This enzyme could be used for the modification of natural fucoidans to obtain more regular and easier characterized derivatives useful for research and practical applications.
authors.affiliation: Laboratory of Enzyme Chemistry, G.B. Elyakov Pacific Institute of Bioorganic Chemistry, Far-Eastern Branch of the Russian Academy of Sciences, 159, Prospect 100-let Vladivostoku, Vladivostok 690022, Russia.
authors.forename: Artem S
authors.initials: AS
authors.lastname: Silchenko
authors.affiliation: Laboratory of Glycoconjugate Chemistry, N.D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences,  47, Leninsky Prospect, Moscow 119991, Russia.
authors.forename: Nadezhda E
authors.initials: NE
authors.lastname: Ustyuzhanina
authors.affiliation: Laboratory of Enzyme Chemistry, G.B. Elyakov Pacific Institute of Bioorganic Chemistry, Far-Eastern Branch of the Russian Academy of Sciences, 159, Prospect 100-let Vladivostoku, Vladivostok 690022, Russia.
authors.forename: Mikhail I
authors.initials: MI
authors.lastname: Kusaykin
authors.affiliation: Laboratory of Glycoconjugate Chemistry, N.D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences,  47, Leninsky Prospect, Moscow 119991, Russia.
authors.forename: Vadim B
authors.initials: VB
authors.lastname: Krylov
authors.affiliation: Laboratory of Glycoconjugate Chemistry, N.D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences,  47, Leninsky Prospect, Moscow 119991, Russia.
authors.forename: Alexander S
authors.initials: AS
authors.lastname: Shashkov
authors.affiliation: Laboratory of Glycoconjugate Chemistry, N.D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences,  47, Leninsky Prospect, Moscow 119991, Russia.
authors.forename: Andrey S
authors.initials: AS
authors.lastname: Dmitrenok
authors.affiliation: Laboratory of Enzyme Chemistry, G.B. Elyakov Pacific Institute of Bioorganic Chemistry, Far-Eastern Branch of the Russian Academy of Sciences, 159, Prospect 100-let Vladivostoku, Vladivostok 690022, Russia.
authors.forename: Roza V
authors.initials: RV
authors.lastname: Usoltseva
authors.affiliation: Far-Eastern Federal University, 8, Sukhanova St., Vladivostok 690022 , Russia.
authors.forename: Anastasiya O
authors.initials: AO
authors.lastname: Zueva
authors.affiliation: Laboratory of Glycoconjugate Chemistry, N.D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences,  47, Leninsky Prospect, Moscow 119991, Russia.
authors.forename: Nikolay E
authors.initials: NE
authors.lastname: Nifantiev
authors.affiliation: Laboratory of Enzyme Chemistry, G.B. Elyakov Pacific Institute of Bioorganic Chemistry, Far-Eastern Branch of the Russian Academy of Sciences, 159, Prospect 100-let Vladivostoku, Vladivostok 690022, Russia.
authors.forename: Tatyana N
authors.initials: TN
authors.lastname: Zvyagintseva
mesh_terms: D003001:Cloning, Molecular; D041965:Flavobacteriaceae / Q000201:enzymology*; D015971:Gene Expression Regulation, Enzymologic; D006026:Glycoside Hydrolases / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D006027:Glycosides / Q000737:chemistry / Q000378:metabolism; D009844:Oligosaccharides / Q000737:chemistry / Q000235:genetics; D011134:Polysaccharides / Q000737:chemistry / Q000378:metabolism*; D011487:Protein Conformation; D013379:Substrate Specificity
chemical_list: D006027:Glycosides; D009844:Oligosaccharides; D011134:Polysaccharides; C007789:fucoidan; D006026:Glycoside Hydrolases
keywords: brown algae; fucoidan; substrate specificity; sulfated oligosaccharides; sulfated polysaccharides