title: Biochemical and Structural Characterization of a Five-domain GH115 α-Glucuronidase from the Marine Bacterium Saccharophagus degradans 2-40T.
abstract: Glucuronic acid (GlcAp) and/or methylglucuronic acid (MeGlcAp) decorate the major forms of xylan in hardwood and coniferous softwoods as well as many cereal grains. Accordingly, the complete utilization of glucuronoxylans or conversion to sugar precursors requires the action of main chain xylanases as well as α-glucuronidases that release the α- (1→2)-linked (Me)GlcAp side groups. Herein, a family GH115 enzymefrom the marine bacterium Saccharophagus degradans 2-40(T), SdeAgu115A, demonstrated activity toward glucuronoxylan and oligomers thereof with preference toward MeGlcAp linked to internal xylopyranosyl residues. Unique biochemical characteristics of NaCl activation were also observed. The crystal structure of SdeAgu115A revealed a five-domain architecture, with an additional insertion C(+) domain that had significant impact on the domain arrangement of SdeAgu115A monomer and its dimerization. The participation of domain C(+) in substrate binding was supported by reduced substrate inhibition upon introducing W773A, W689A, and F696A substitutions within this domain. In addition to Asp-335, the catalytic essentiality of Glu-216 was revealed by site-specific mutagenesis. A primary sequence analysis suggested that the SdeAgu115A architecture is shared by more than half of GH115 members, thus defining a distinct archetype for GH115 enzymes.
authors.affiliation: Department of Chemical Engineering and Applied Chemistry, University of Toronto, 200 College Street, Toronto, Ontario M5S 3E5, Canada.
authors.forename: Weijun
authors.initials: W
authors.lastname: Wang
authors.affiliation: Department of Chemical Engineering and Applied Chemistry, University of Toronto, 200 College Street, Toronto, Ontario M5S 3E5, Canada.
authors.forename: Ruoyu
authors.initials: R
authors.lastname: Yan
authors.affiliation: Structural Biology Center, Biosciences Division, Argonne National Laboratory, Argonne, Illinois 60439.
authors.forename: Boguslaw P
authors.initials: BP
authors.lastname: Nocek
authors.affiliation: Department of Chemical Engineering and Applied Chemistry, University of Toronto, 200 College Street, Toronto, Ontario M5S 3E5, Canada.
authors.forename: Thu V
authors.initials: TV
authors.lastname: Vuong
authors.affiliation: Department of Chemical Engineering and Applied Chemistry, University of Toronto, 200 College Street, Toronto, Ontario M5S 3E5, Canada.
authors.forename: Rosa
authors.initials: R
authors.lastname: Di Leo
authors.affiliation: Department of Chemical Engineering and Applied Chemistry, University of Toronto, 200 College Street, Toronto, Ontario M5S 3E5, Canada.
authors.forename: Xiaohui
authors.initials: X
authors.lastname: Xu
authors.affiliation: Department of Chemical Engineering and Applied Chemistry, University of Toronto, 200 College Street, Toronto, Ontario M5S 3E5, Canada.
authors.forename: Hong
authors.initials: H
authors.lastname: Cui
authors.affiliation: Department of Chemistry and Chemical Engineering, Wallenberg Wood Science Center and Biopolymer Technology, Chalmers University of Technology, Kemivägen 4, Gothenburg 412 96, Sweden.
authors.forename: Paul
authors.initials: P
authors.lastname: Gatenholm
authors.affiliation: Department of Chemistry and Chemical Engineering, Wallenberg Wood Science Center and Biopolymer Technology, Chalmers University of Technology, Kemivägen 4, Gothenburg 412 96, Sweden,; Department of Wood, Cellulose and Paper Research, University of Guadalajara, Guadalajara 44100, Mexico.
authors.forename: Guillermo
authors.initials: G
authors.lastname: Toriz
authors.affiliation: Department of Food and Environmental Sciences, University of Helsinki, P.O. Box 27, Helsinki 00014, Finland.
authors.forename: Maija
authors.initials: M
authors.lastname: Tenkanen
authors.affiliation: Department of Chemical Engineering and Applied Chemistry, University of Toronto, 200 College Street, Toronto, Ontario M5S 3E5, Canada,. Electronic address: alexei.savchenko@utoronto.ca.
authors.forename: Alexei
authors.initials: A
authors.lastname: Savchenko
authors.affiliation: Department of Chemical Engineering and Applied Chemistry, University of Toronto, 200 College Street, Toronto, Ontario M5S 3E5, Canada,. Electronic address: emma.master@utoronto.ca.
authors.forename: Emma R
authors.initials: ER
authors.lastname: Master
mesh_terms: D000595:Amino Acid Sequence; D002942:Circular Dichroism; D020563:Gammaproteobacteria / Q000201:enzymology*; D006026:Glycoside Hydrolases / Q000737:chemistry / Q000378:metabolism*; D008386:Marine Biology; D008958:Models, Molecular; D011487:Protein Conformation; D017386:Sequence Homology, Amino Acid
chemical_list: D006026:Glycoside Hydrolases; C096734:alpha-glucuronidase
keywords: GH115 α-glucuronidase; carbohydrate; carbohydrate degradation; catalytic apparatus; crystal structure; enzyme catalysis; enzyme domain composition; enzyme structure; protein domain