BioMetals 2009-12-01

Biochemical characterization of Helicobacter pylori α-1,4 fucosyltransferase: metal ion requirement, donor substrate specificity and organic solvent stability.

Said Rabbani, Francesco Corona, Beat Ernst

Index: Biometals 22(6) , 1011-7, (2009)

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Abstract

The effect of metal ions on the activity, the donor substrate specificity, and the stability in organic solvents of Helicobacter pylori α-1,4 fucosyltransferase were studied. The recombinant enzyme was expressed as soluble form in E. coli strain AD494 and purified in a one step affinity chromatography. Its activity was highest in cacodylate buffer at pH 6.5 in the presence of 20 mM Mn2+ ions at 37°C. Mn2+ ions could be substituted by other metal ions. In all cases, Mn2+ ions proofed to be the most effective (Mn2+ > Co2+ > Ca2+ > Mg2+ > Cu2+ > Ni2+ > EDTA). The enzyme shows substrate specificity for Type I disaccharide (1) with a KM of 114 μM. In addition, the H. pylori α-1,4 fucosyltransferase efficiently transfers GDP-activated L-fucose derivatives to Galβ1-3GlcNAc-OR (1). Interestingly, the presence of organic solvents such as DMSO and methanol up to 20% in the reaction medium does not affect significantly the enzyme activity. However, at the same concentration of dioxane, activity is totally abolished.