title: Crystal structure of α-1,4-glucan lyase, a unique glycoside hydrolase family member with a novel catalytic mechanism.
abstract: α-1,4-Glucan lyase (EC 4.2.2.13) from the red seaweed Gracilariopsis lemaneiformis cleaves α-1,4-glucosidic linkages in glycogen, starch, and malto-oligosaccharides, yielding the keto-monosaccharide 1,5-anhydro-D-fructose. The enzyme belongs to glycoside hydrolase family 31 (GH31) but degrades starch via an elimination reaction instead of hydrolysis. The crystal structure shows that the enzyme, like GH31 hydrolases, contains a (β/α)8-barrel catalytic domain with B and B' subdomains, an N-terminal domain N, and the C-terminal domains C and D. The N-terminal domain N of the lyase was found to bind a trisaccharide. Complexes of the enzyme with acarbose and 1-dexoynojirimycin and two different covalent glycosyl-enzyme intermediates obtained with fluorinated sugar analogues show that, like GH31 hydrolases, the aspartic acid residues Asp(553) and Asp(665) are the catalytic nucleophile and acid, respectively. However, as a unique feature, the catalytic nucleophile is in a position to act also as a base that abstracts a proton from the C2 carbon atom of the covalently bound subsite -1 glucosyl residue, thus explaining the unique lyase activity of the enzyme. One Glu to Val mutation in the active site of the homologous α-glucosidase from Sulfolobus solfataricus resulted in a shift from hydrolytic to lyase activity, demonstrating that a subtle amino acid difference can promote lyase activity in a GH31 hydrolase.
authors.affiliation: From the Laboratory of Biophysical Chemistry, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Nijenborgh 7, 9747 AG Groningen, The Netherlands.
authors.forename: Henriëtte J
authors.initials: HJ
authors.lastname: Rozeboom
authors.forename: Shukun
authors.initials: S
authors.lastname: Yu
authors.forename: Susan
authors.initials: S
authors.lastname: Madrid
authors.forename: Kor H
authors.initials: KH
authors.lastname: Kalk
authors.forename: Ran
authors.initials: R
authors.lastname: Zhang
authors.forename: Bauke W
authors.initials: BW
authors.lastname: Dijkstra
mesh_terms: D020909:Acarbose / Q000737:chemistry; D002384:Catalysis; D020134:Catalytic Domain; D002851:Chromatography, High Pressure Liquid; D018360:Crystallography, X-Ray; D005936:Glucans / Q000737:chemistry; D018698:Glutamic Acid / Q000737:chemistry; D006026:Glycoside Hydrolases / Q000737:chemistry*; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D011133:Polysaccharide-Lyases / Q000737:chemistry*; D017434:Protein Structure, Tertiary; D011522:Protons; D012624:Seaweed / Q000201:enzymology*; D048229:Sulfolobus solfataricus / Q000201:enzymology; D014312:Trisaccharides / Q000737:chemistry; D014633:Valine / Q000737:chemistry; D000520:alpha-Glucosidases / Q000378:metabolism
chemical_list: D005936:Glucans; D011522:Protons; D014312:Trisaccharides; D018698:Glutamic Acid; D006026:Glycoside Hydrolases; D000520:alpha-Glucosidases; D011133:Polysaccharide-Lyases; C080474:alpha-1,4-glucan lyase; D014633:Valine; D020909:Acarbose
keywords: 1-Deoxynojirimycin; Acarbose; Anhydrofructose Pathway; Carbohydrate Binding Site; Crystal Structure; Enzyme Mechanisms; Glycoside Hydrolases; Glycosylation Inhibitors; Glycosylenzyme Intermediate; X-ray Crystallography
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