Characterization of recombinant UDP-galactopyranose mutase from Aspergillus fumigatus

UDP-galactopyranose mutase (UGM) is a flavin-containing enzyme that catalyzes the conversion of UDP-galactopyranose to UDP-galactofuranose, the precursor of galactofuranose, which is an important cell wall component in Aspergillus fumigatus and other pathogenic microbes. A. fumigatus UGM (AfUGM) was expressed in Escherichia coli and purified to homogeneity. The enzyme was shown to function as a homotetramer by size-exclusion chromatography and to contain ∼50% of the flavin in the active reduced form. A kcat value of 72 ± 4 s−1 and a KM value of 110 ± 15 μM were determined with UDP-galactofuranose as substrate. In the oxidized state, AfUGM does not bind UDP-galactopyranose, while UDP and UDP-glucose bind with Kd values of 33 ± 9 μM and 90 ± 30 μM, respectively. Functional and structural differences between the bacterial and eukaryotic UGMs are discussed.

Research highlights
► Recombinant UDP-galactopyranose (AfUGM) from A. fumigatus.
► AfUGM is a tetrameric enzyme.
► Under aerobic conditions, half of the flavin is present in the reduced active form.