Crystallization and X-ray diffraction studies of cellobiose phosphorylase from Cellulomonas uda.
Disaccharide phosphorylases are able to catalyze both the synthesis and the breakdown of disaccharides and have thus emerged as attractive platforms for tailor-made sugar synthesis. Cellobiose phosphorylase from Cellulomonas uda (CPCuda) is an enzyme that belongs to glycoside hydrolase family 94 and catalyzes the reversible breakdown of cellobiose [beta-D-glucopyranosyl-(1,4)-D-glucopyranose] to alpha-D-glucose-1-phosphate and D-glucose. Crystals of ligand-free recombinant CPCuda and of its complexes with substrates and reaction products yielded complete X-ray diffraction data sets to high resolution using synchrotron radiation but suffered from significant variability in diffraction quality. In at least one case an intriguing space-group transition from a primitive monoclinic to a primitive orthorhombic lattice was observed during data collection. The structure of CPCuda was determined by maximum-likelihood molecular replacement, thus establishing a starting point for an investigation of the structural and mechanistic determinants of disaccharide phosphorylase activity.
Van Hoorebeke
Annelies
A
Unit for Structural Biology and Biophysics, Laboratory for Protein Biochemistry and Biomolecular Engineering, K. L. Ledeganckstraat 35, Ghent University, B-9000 Ghent, Belgium.
Stout
Jan
J
Kyndt
John
J
De Groeve
Manu
M
Dix
Ina
I
Desmet
Tom
T
Soetaert
Wim
W
Van Beeumen
Jozef
J
Savvides
Savvas N
SN
D040143:Cellulomonas / Q000201:enzymology*; D003460:Crystallization; D018360:Crystallography, X-Ray; D005964:Glucosyltransferases / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D008958:Models, Molecular; D020836:Protein Structure, Quaternary; D013379:Substrate Specificity
D005964:Glucosyltransferases; C034714:cellobiose phosphorylase