Crystallization and X-ray diffraction studies of cellobiose phosphorylase from Cellulomonas uda

• Published in: Acta crystallographica. Section F, Structural biology and crystallization communications Structural biology and crystallization communications, 2010-03, Vol.66 (3), p.346-351
• Main Authors: Van Hoorebeke, Annelies, Stout, Jan, Kyndt, John, De Groeve, Manu, Dix, Ina, Desmet, Tom, Soetaert, Wim, Van Beeumen, Jozef, Savvides, Savvas N.
• Format: Article
• Language: English
• Subjects: cellobiose phosphorylases; Cellulomonas - enzymology; Crystallization; Crystallization Communications; Crystallography, X-Ray; disaccharide phosphorylases; GH94; Glucosyltransferases - chemistry; Glucosyltransferases - isolation & purification; Glucosyltransferases - metabolism; inverting hydrolases; Models, Molecular; Protein Structure, Quaternary; Substrate Specificity
• ISSN: 1744-3091; 1744-3091; 2053-230X
• DOI: 10.1107/S1744309110002642

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 [β‐d‐glucopyranosyl‐(1,4)‐d‐glucopyranose] to α‐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.