Many pathogenic prokaryotes and eukaryotes possess the machinery required to assemble galactofuranose (Galf)-containing glycoconjugates; these glycoconjugates can be critical for virulence or viability. Accordingly, compounds that block Galf incorporation may serve as therapeutic leads or as probes of the function of Galf-containing glycoconjugates. The enzyme UDP-galactopyranose mutase (UGM) is the only known generator of UDP-galactofuranose, the precursor to Galf residues. We previously employed a high-throughput fluorescence polarization assay to investigate the Klebsiella pneumoniae UGM. We demonstrate the generality of this assay by extending it to UGM from Mycobacterium tuberculosis. To identify factors influencing binding, we synthesized a directed library containing a 5-arylidene-2-thioxo-4-thiazolidinone core, a structure possessing features common to ligands for both homologs. Our studies offer a blueprint for identifying inhibitors of the growing family of UGM homologs and provide insight into UGM inhibition.
Bibliographical noteFunding Information:
We thank M. Soltero-Higgin for determining the K d and IC 50 values of compounds 5 , 6 , 7 , and 8 with UGM Kleb , for determining the K d value of UDP with UGM Kleb , and for performing initial experiments to produce UGM Myco . We also thank J. Phillips for the synthesis of compound 1 and A. Steinberg for assistance in generating Figure 5 . We acknowledge N. Peters, M. Fitzgerald, and M. Hoffmann of the Keck-UWCCC Small Molecule Screening Facility for helpful discussions. MALDI-TOF MS data were obtained at the University of Wisconsin-Madison Biophysics Instrumentation Facility, which is supported by the University of Wisconsin-Madison, the National Science Foundation (BIR-9512577), and the National Institutes of Health (RR13790). This research was supported by the National Institutes of Health (AI635976). E.E.C. was supported by the National Institutes of Health Biotechnology Training Program (GM08349), and J.F.M. was supported by a National Science Foundation Graduate Research Fellowship and a Wisconsin Alumni Research Foundation Fellowship.