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4D48: Crystal Structure of glucose-1-phosphate uridylyltransferase GalU from Erwinia amylovora
Glucose-1-phosphate uridylyltransferase from Erwinia amylovora: Activity, structure and substrate specificity
Biochim. Biophys. Acta (2017) 1865 p.1348-1357» All references (2)
Erwinia amylovora, a Gram-negative plant pathogen, is the causal agent of Fire Blight, a contagious necrotic disease affecting plants belonging to the Rosaceae family, including apple and pear. E. amylovora is highly virulent and capable of rapid dissemination in orchards; effective control methods are still lacking. One of its most important pathogenicity factors is the exopolysaccharide amylovoran. Amylovoran is a branched polymer made by the repetition of units mainly composed of galactose, with some residues of glucose, glucuronic acid and pyruvate. E. amylovora glucose-1-phosphate uridylyltransferase (UDP-glucose pyrophosphorylase, EC 22.214.171.124) has a key role in amylovoran biosynthesis. This enzyme catalyses the production of UDP-glucose from glucose-1-phosphate and UTP, which the epimerase GalE converts into UDP-galactose, the main building block of amylovoran. We determined EaGalU kinetic parameters and substrate specificity with a range of sugar 1-phosphates. At time point 120min the enzyme catalysed conversion of the sugar 1-phosphate into the corresponding UDP-sugar reached 74% for N-acetyl-alpha-d-glucosamine 1-phosphate, 28% for alpha-d-galactose 1-phosphate, 0% for alpha-d-galactosamine 1-phosphate, 100% for alpha-d-xylose 1-phosphate, 100% for alpha-d-glucosamine 1-phosphate, 70% for alpha-d-mannose 1-phosphate, and 0% for alpha-d-galacturonic acid 1-phosphate. To explain our results we obtained the crystal structure of EaGalU and augmented our study by docking the different sugar 1-phosphates into EaGalU active site, providing both reliable models for substrate binding and enzyme specificity, and a rationale that explains the different activity of EaGalU on the sugar 1-phosphates used. These data demonstrate EaGalU potential as a biocatalyst for biotechnological purposes, as an alternative to the enzyme from Escherichia coli, besides playing an important role in E. amylovora pathogenicity.