title: A new archaeal beta-glycosidase from Sulfolobus solfataricus: seeding a novel retaining beta-glycan-specific glycoside hydrolase family along with the human non-lysosomal glucosylceramidase GBA2.
abstract: Carbohydrate active enzymes (CAZymes) are a large class of enzymes, which build and breakdown the complex carbohydrates of the cell. On the basis of their amino acid sequences they are classified in families and clans that show conserved catalytic mechanism, structure, and active site residues, but may vary in substrate specificity. We report here the identification and the detailed molecular characterization of a novel glycoside hydrolase encoded from the gene sso1353 of the hyperthermophilic archaeon Sulfolobus solfataricus. This enzyme hydrolyzes aryl beta-gluco- and beta-xylosides and the observation of transxylosylation reactions products demonstrates that SSO1353 operates via a retaining reaction mechanism. The catalytic nucleophile (Glu-335) was identified through trapping of the 2-deoxy-2-fluoroglucosyl enzyme intermediate and subsequent peptide mapping, while the general acid/base was identified as Asp-462 through detailed mechanistic analysis of a mutant at that position, including azide rescue experiments. SSO1353 has detectable homologs of unknown specificity among Archaea, Bacteria, and Eukarya and shows distant similarity to the non-lysosomal bile acid beta-glucosidase GBA2 also known as glucocerebrosidase. On the basis of our findings we propose that SSO1353 and its homologs are classified in a new CAZy family, named GH116, which so far includes beta-glucosidases (EC 3.2.1.21), beta-xylosidases (EC 3.2.1.37), and glucocerebrosidases (EC 3.2.1.45) as known enzyme activities.
authors.affiliation: Institute of Protein Biochemistry, CNR, Via P. Castellino 111, 80131, Naples, Italy.
authors.forename: Beatrice
authors.initials: B
authors.lastname: Cobucci-Ponzano
authors.forename: Vincenzo
authors.initials: V
authors.lastname: Aurilia
authors.forename: Gennaro
authors.initials: G
authors.lastname: Riccio
authors.forename: Bernard
authors.initials: B
authors.lastname: Henrissat
authors.forename: Pedro M
authors.initials: PM
authors.lastname: Coutinho
authors.forename: Andrea
authors.initials: A
authors.lastname: Strazzulli
authors.forename: Anna
authors.initials: A
authors.lastname: Padula
authors.forename: Maria Michela
authors.initials: MM
authors.lastname: Corsaro
authors.forename: Giuseppina
authors.initials: G
authors.lastname: Pieretti
authors.forename: Gabriella
authors.initials: G
authors.lastname: Pocsfalvi
authors.forename: Immacolata
authors.initials: I
authors.lastname: Fiume
authors.forename: Raffaele
authors.initials: R
authors.lastname: Cannio
authors.forename: Mosè
authors.initials: M
authors.lastname: Rossi
authors.forename: Marco
authors.initials: M
authors.lastname: Moracci
mesh_terms: D017931:DNA Primers; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D005784:Gene Amplification; D005959:Glucosidases / Q000145:classification / Q000235:genetics* / Q000378:metabolism*; D005962:Glucosylceramidase / Q000145:classification / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D009844:Oligosaccharides / Q000494:pharmacology; D010802:Phylogeny; D011994:Recombinant Proteins / Q000302:isolation & purification / Q000378:metabolism; D021241:Spectrometry, Mass, Electrospray Ionization; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013379:Substrate Specificity; D016944:Sulfolobus / Q000201:enzymology; D014995:Xylosidases / Q000145:classification / Q000378:metabolism; D001617:beta-Glucosidase / Q000145:classification / Q000378:metabolism*
chemical_list: D017931:DNA Primers; D009844:Oligosaccharides; D011994:Recombinant Proteins; D005959:Glucosidases; C104851:Sulfolobus solfataricus beta-glycosidase; D014995:Xylosidases; D001617:beta-Glucosidase; C026579:exo-1,4-beta-D-xylosidase; C517537:GBA2 protein, human; D005962:Glucosylceramidase