title: Specificity of Processing α-glucosidase I is guided by the substrate conformation: crystallographic and in silico studies.
abstract: BACKGROUND
  The enzyme “GluI” is key to the synthesis of critical glycoproteins in the cell.
  
  
  RESULTS
  We have determined the structure of GluI, and modeled binding with its unique sugar substrate.
  
  
  CONCLUSIONS
  The specificity of this interaction derives from a unique conformation of the substrate.
  
  
  CONCLUSIONS
  Understanding the mechanism of the enzyme is of basic importance and relevant to potential development of antiviral inhibitors. Processing α-glucosidase I (GluI) is a key member of the eukaryotic N-glycosylation processing pathway, selectively catalyzing the first glycoprotein trimming step in the endoplasmic reticulum. Inhibition of GluI activity impacts the infectivity of enveloped viruses; however, despite interest in this protein from a structural, enzymatic, and therapeutic standpoint, little is known about its structure and enzymatic mechanism in catalysis of the unique glycan substrate Glc3Man9GlcNAc2. The first structural model of eukaryotic GluI is here presented at 2-Å resolution. Two catalytic residues are proposed, mutations of which result in catalytically inactive, properly folded protein. Using Autodocking methods with the known substrate and inhibitors as ligands, including a novel inhibitor characterized in this work, the active site of GluI was mapped. From these results, a model of substrate binding has been formulated, which is most likely conserved in mammalian GluI.
authors.affiliation: Department of Medical Biophysics, University of Toronto, Ontario Cancer Institute, Princess Margaret Hospital, Toronto, Ontario M5G 2M9, Canada. megan.barker@utoronto.ca
authors.forename: Megan K
authors.initials: MK
authors.lastname: Barker
authors.forename: David R
authors.initials: DR
authors.lastname: Rose
mesh_terms: D017485:1-Deoxynojirimycin / Q000031:analogs & derivatives / Q000737:chemistry; D002236:Carbohydrate Conformation; D002240:Carbohydrate Sequence; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D005947:Glucose / Q000737:chemistry; D065089:Glycoside Hydrolase Inhibitors; D006860:Hydrogen Bonding; D007700:Kinetics; D008562:Membrane Glycoproteins / Q000037:antagonists & inhibitors / Q000737:chemistry*; D062105:Molecular Docking Simulation; D008969:Molecular Sequence Data; D011485:Protein Binding; D017433:Protein Structure, Secondary; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D029701:Saccharomyces cerevisiae Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry*; D040681:Structural Homology, Protein; D013379:Substrate Specificity; D014312:Trisaccharides / Q000737:chemistry; D000520:alpha-Glucosidases / Q000737:chemistry*
chemical_list: D065089:Glycoside Hydrolase Inhibitors; D008562:Membrane Glycoproteins; D029701:Saccharomyces cerevisiae Proteins; D014312:Trisaccharides; C045621:miglitol; D017485:1-Deoxynojirimycin; C098060:CWH41 protein, S cerevisiae; D000520:alpha-Glucosidases; D005947:Glucose
keywords: Carbohydrate Glycoprotein; Computer Modeling; Docking; Enzyme Structure; Glycobiology; Glycosylation; Post-translational Modification; X-ray Crystallography
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