UDP-Galactopyranose Mutase in Nematodes

Abstract

Nematodes represent a diverse phylum of both free living and parasitic species. While the species Caenorhabditis elegans (C. elegans) is a valuable model organism, parasitic nematodes or helminths pose a serious threat to human health. Indeed, helminths cause many neglected tropical diseases that afflict humans. Nematode glycoconjugates have been implicated in evasive immunomodulation, a hallmark of nematode infections. One monosaccharide residue present in the glycoconjugates of several human pathogens is galactofuranose (Galf). This five-membered ring isomer of galactose has not been detected in mammals, making Galf metabolic enzymes attractive therapeutic targets. The only known pathway for biosynthetic incorporation of Galf into glycoconjugates depends upon generation of the glycosyl donor UDP-Galf by the flavoenzyme uridine 5’-diphosphate (UDP) galactopyranose mutase (UGM or Glf). A putative UGM encoding gene (glf-1) was recently identified in C. elegans. Because Galf has yet to be identified in any nematode glycan, we sought to assess the catalytic activity of the C. elegans glf-1 gene product (CeUGM). We found that CeUGM catalyzes the isomerization of UDP-Galf and UDP-galactopyranose (UDP-Galp). In the presence of enzyme, substrate, and a hydride source, a galactose–N5-FAD adduct was isolated, suggesting the CeUGM flavin adenine dinucleotide (FAD) cofactor serves as a nucleophile in covalent catalysis. Homology modeling and protein variants indicate that CeUGM possesses an active site similar to that of prokaryotic enzymes, despite the low sequence identity (~15%) between eukaryotic and prokaryotic UGM proteins. Even with the primary sequence differences, heterocyclic UGM inhibitors developed against prokaryotic proteins also inhibit CeUGM activity. We postulate that these inhibitors can serve as chemical probes of Galf in nematodes and as anthelmintic leads. Together, our data suggest that CeUGM facilitates the biosynthetic incorporation of Galf into nematode glycoconjugates through generation of the glycosyl donor UDP-Galf.

UDP-galactopyranose mutases from Leishmania species that cause visceral and cutaneous leishmaniasis

“…3. Comparison with available data from the other eukaryotic UGMs show that, although there are minor differences between the k cat and K m values, the catalytic efficiencies ( k cat / K m ) are very similar among all eukaryotic enzymes, with the exception of A. fumigatus UGM (Table 1) [18, 19, 27]. …”

“…It has been shown that in the reduced form, the FAD-N5 attacks the UDP-Gal p at the C1 position, leading to breaking of the anomeric bond and formation of an FAD-galactose adduct. This adduct is essential for sugar ring contraction [9, 14, 27, 31]. Recently, we showed that Tc UGM and Af UGM can be reduced by NADPH and to a lesser extent by NADH [14].…”

Bioinformatics and Drug Discovery

“…Because Gal f is absent in human [44], UGM has been used as a desirable drug target [49]. UGM coded by gene GLF was later found in several eukaryotic unicellular pathogens [50] as well as in nematodes [51]. Can we repurposing drugs developed against bacterial pathogens to fight eukaryotic unicellular pathogens [50]?…”

Conformational Control of UDP-Galactopyranose Mutase Inhibition

“…7, 8 Although Gal f -containing glycans in nematodes have yet to be identified, nematodes encode a UGM, and Caenorhabditis elegans expresses a functional UGM. 9-11 In contrast, Gal f residues are not found in mammals nor do mammals possess UGM. 12 In many organisms that encode a UGM, deletion or downregulation of UGM production has deleterious consequences, 9, 13 including lethality in mycobacteria.…”