Phosphoglucomutase is absent in Trypanosoma brucei and redundantly substituted by phosphomannomutase and phospho‐N‐acetylglucosamine mutase

Abstract

The enzymes phosphomannomutase (PMM), phospho-N-acetylglucosamine mutase (PAGM) and phosphoglucomutase (PGM) reversibly catalyse the transfer of phosphate between the C6 and C1 hydroxyl groups of mannose, N-acetylglucosamine and glucose respectively. Although genes for a candidate PMM and a PAGM enzymes have been found in the Trypanosoma brucei genome, there is, surprisingly, no candidate gene for PGM. The TbPMM and TbPAGM genes were cloned and expressed in Escherichia coli and the TbPMM enzyme was crystallized and its structure solved at 1.85 Å resolution. Antibodies to the recombinant proteins localized endogenous TbPMM to glycosomes in the bloodstream form of the parasite, while TbPAGM localized to both the cytosol and glycosomes. Both recombinant enzymes were able to interconvert glucose-phosphates, as well as acting on their own definitive substrates. Analysis of sugar nucleotide levels in parasites with TbPMM or TbPAGM knocked down by RNA interference (RNAi) suggests that, in vivo, PGM activity is catalysed by both enzymes. This is the first example in any organism of PGM activity being completely replaced in this way and it explains why, uniquely, T. brucei has been able to lose its PGM gene. The RNAi data for TbPMM also showed that this is an essential gene for parasite growth.

Biology, Mechanism, and Structure of Enzymes in the α- d -Phosphohexomutase Superfamily

“…This can make them more difficult to identify as a member of the superfamily based on sequence searches. The sequence motif of the sugar-binding loop in the PAGMs is distinct: –FEANG– (Bandini et al, 2012; Kato et al, 2005) (Table 1 and Fig. 2), with the asparagine reflecting a role in recognizing the N -acetyl group of the 1-phospho form of the sugar (Nishitani et al, 2006).…”

Comparative Metabolism of Free‐living Bodo saltans and Parasitic Trypanosomatids

“…Indeed, it was demonstrated that PAGM is capable of replacing PGM in T. brucei (Bandini et al. ). Galactose is essential for T. brucei , but the parasite not only lacks transporters for its uptake, but also a gene for galactokinase.…”

“…PGM may be replaced by phospho-N-acetylglucosamine mutase (PAGM), which reversibly catalyzes the transfer of phosphate between the C6 and C1 hydroxyl groups of Nacetylglucosamine, mannose, glucose, and phospho-Nacetylglucosamine. Indeed, it was demonstrated that PAGM is capable of replacing PGM in T. brucei (Bandini et al 2012). Galactose is essential for T. brucei, but the parasite not only lacks transporters for its uptake, but also a gene for galactokinase.…”

Proteins and lipids of glycosomal membranes from Leishmania tarentolae and Trypanosoma brucei

“…These additional enzymes involved in the metabolism of sugars might indicate a higher metabolic flexibility of L. tarentolae compared to African trypanosomes. In T. brucei , the phosphomannomutase Tb927.10.6440 was found in the glycosome, where it can act as phosphoglucomutase during glycolysis 36 . T. brucei phospho-N-acetylglucosamine mutase (Tb927.8.980) was also partially glycosomal 36 .…”

“…In T. brucei , the phosphomannomutase Tb927.10.6440 was found in the glycosome, where it can act as phosphoglucomutase during glycolysis 36 . T. brucei phospho-N-acetylglucosamine mutase (Tb927.8.980) was also partially glycosomal 36 . Neither has an obvious PTS1 targeting signal so it is possible that they have either an internal glycosomal targeting signal or that they are co-imported via association with other glycosomal targeting signal-containing proteins 37, 38 .…”