Mechanistic studies on UDP‐Galactopyranose mutases from Aspergillus fumigatus and Trypanosoma cruzi

Abstract

UDP‐Galactopyranose mutase (UGM) is a flavoprotein that catalyzes the conversion of UDP‐galactopyranose to UDP‐galactofuranose. UGM serves as the sole biosynthetic source of galactofuranose, which is found on the cell surface of many bacteria, fungi, and parasites. UGM is a unique flavoprotein since it requires the flavin to be reduced for activity, yet no net redox change occurs during catalysis. Currently, the exact mechanism of isomerization is unknown. UGM is also of interest because it is a novel drug target for several diseases due to the importance of galactofuranose for the survival of M. tuberculosis and for virulence of the fungus, A. fumigatus, and the human parasites, such as L. major and T. cruzi. While many studies have been done on the prokaryotic UGM, little is known about eukaryotic UGM. We present, for the first time, the recombinant expression, purification, and characterization of eukaryotic UGMs from A. fumigatus and T. cruzi. We also report limited proteolysis, steady‐state kinetics, rapid‐reaction kinetics, and site‐directed mutagenesis data that demonstrate significant differences between prokaryotic and eukaryotic UGMs.