Crystallization and preliminary X-ray analysis of the class 1 alpha 1,2-mannosidase from Saccharomyces cerevisiae.
The alpha 1,2-mannosidase from Saccharomyces cerevisiae catalyzes the conversion of Man9GlcNAc2 to Man8GlcNAc2 during the formation of N-linked oligosaccharides and is a member of the Class 1 alpha 1,2-mannosidases conserved from yeast to mammals. The enzyme is a type II membrane protein and a recombinant form of the alpha 1,2-mannosidase from S. cerevisiae, lacking the transmembrane domain, has been expressed in Pichia pastoris and crystallized using the hanging drop vapor diffusion technique. The crystals grow as flat plates, with unit cell dimensions a = 57.5 A, b = 84.1 A, c = 107.1 A, alpha = beta = gamma = 90 degrees. The crystals exhibit the symmetry of space group P2(1)2(1)2(1) and diffract to a minimum d-spacing of 3.5 A resolution. On the basis of density calculations one monomer is estimated to be present in the asymmetric unit (Vm = 2.08 A3 Da-1). This is the first report of the crystallization of any glycosidase involved in N-glycan biosynthesis.
Dole
K
K
Division of Biochemistry Research, Hospital for Sick Children, Toronto, Ontario, Canada.
Lipari
F
F
Herscovics
A
A
Howell
P L
PL
D002240:Carbohydrate Sequence; D018360:Crystallography, X-Ray / Q000379:methods; D008361:Mannosidases / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D008565:Membrane Proteins / Q000737:chemistry / Q000302:isolation & purification; D008969:Molecular Sequence Data; D011134:Polysaccharides / Q000096:biosynthesis / Q000737:chemistry; D011994:Recombinant Proteins / Q000737:chemistry / Q000302:isolation & purification; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D013379:Substrate Specificity
D008565:Membrane Proteins; D011134:Polysaccharides; D011994:Recombinant Proteins; D008361:Mannosidases; C046090:mannosyl-oligosaccharide 1,2-alpha-mannosidase