title: Refined structure for the complex of 1-deoxynojirimycin with glucoamylase from Aspergillus awamori var. X100 to 2.4-A resolution.
abstract: The three-dimensional structure of the complex of 1-deoxynojirimycin with glucoamylase II-(471) from Aspergillus awamori var. X100 has been determined to 2.4-A resolution. The model includes residues corresponding to residues 1-471 of glucoamylase I from Aspergillus niger, two molecules of bound 1-deoxynojirimycin and 605 sites for water molecules. The crystallographic R factor from refinement is 0.119, and the root-mean-squared deviation in bond distances is 0.012 A. The inhibitor complex confirms the location of the active site in the packing void of the alpha/alpha-barrel as proposed by Aleshin et al. [Aleshin, A., Golubev, A., Firsov, L., & Honzatko, R. B. (1992) J. Biol. Chem. 267, 19291-19298]. One inhibitor molecule is associated with strong electron density and represents the principal site of interaction of 1-deoxynojirimycin with the enzyme. The other 1-deoxynojirimycin molecule is associated with weak electron density and therefore, probably represents a binding site of low affinity. Interactions of 1-deoxynojirimycin with the enzyme at its principal site involve Arg 45, Asp 55, Arg 305, and carbonyl 177. In addition, a water molecule (water 500) hydrogen bonds to Glu 400 and the 6-hydroxyl of 1-deoxynojirimycin and is at an approximate distance of 3.3 A from the "anomeric" carbon of the inhibitor. The structural arrangement of functional groups near the inhibitor molecule suggests that Glu 179 is a catalytic acid, Glu 400 a catalytic base, and water 500 the attacking nucleophile in the hydrolysis of maltooligosaccharides. The relevance of the X-ray work to proposed mechanisms of enzymatic hydrolysis of oligosaccharides is discussed.
authors.affiliation: Department of Biochemistry and Biophysics, Iowa State University, Ames 50011.
authors.forename: E M
authors.initials: EM
authors.lastname: Harris
authors.forename: A E
authors.initials: AE
authors.lastname: Aleshin
authors.forename: L M
authors.initials: LM
authors.lastname: Firsov
authors.forename: R B
authors.initials: RB
authors.lastname: Honzatko
mesh_terms: D017485:1-Deoxynojirimycin / Q000737:chemistry / Q000378:metabolism*; D000595:Amino Acid Sequence; D001230:Aspergillus / Q000201:enzymology*; D001665:Binding Sites; D005087:Glucan 1,4-alpha-Glucosidase / Q000737:chemistry* / Q000378:metabolism*; D006860:Hydrogen Bonding; D008958:Models, Molecular; D008968:Molecular Conformation; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000378:metabolism; D017433:Protein Structure, Secondary; D014961:X-Ray Diffraction / Q000379:methods
chemical_list: D009844:Oligosaccharides; D017485:1-Deoxynojirimycin; D005087:Glucan 1,4-alpha-Glucosidase