Abstract
Trypanosoma brucei, a protist parasite that causes African trypanosomiasis or sleeping sickness, relies mainly on glycolysis for ATP production when in its mammalian host. Glycolysis occurs within a peroxisome-like organelle named the glycosome. Previous work from our laboratory reported the presence of significant amounts of inorganic polyphosphate (polyP), a polymer of three to hundreds of orthophosphate units, in the glycosomes and nucleoli of T. brucei. In this work, we identified and characterized the activity of two Nudix hydrolases (NHs), T. brucei Nudix hydrolase (TbNH) 2 and TbNH4, one located in the glycosomes and the other in the cytosol and nucleus, respectively, which can degrade polyP. We found that TbNH2 is an exopolyphosphatase with higher activity on short chain polyP, while TbNH4 is an endo- and exopolyphosphatase that has similar activity on polyP of various chain sizes. Both enzymes have higher activity at around pH 8.0. We also found that only TbNH2 can dephosphorylate ATP and ADP but with lower affinity than for polyP. Our results suggest that NHs can participate in polyP homeostasis and therefore may help control polyP levels in glycosomes, cytosol and nuclei of T. brucei.
Highlights
Inorganic polyphosphate is a linear polymer of phosphate that can range from three to hundreds of Pi units
In Trypanosoma brucei, one of the agents of African trypanosomiasis, polyP is synthesized by the vacuolar transporter chaperone (VTC) complex [2], which is located in the acidocalcisome [3], an acidic organelle that stores calcium and other cations together with Pi, inorganic pyrophosphate (PPi), and polyP [4]
The most important findings of this work are the identification of two nudix hydrolase (NH) of T. brucei as polyP phosphatases, and evidence of their localization in subcellular organelles where polyP is or may be present
Summary
Inorganic polyphosphate (polyP) is a linear polymer of phosphate (orthophosphate [Pi]) that can range from three to hundreds of Pi units. In Trypanosoma brucei, one of the agents of African trypanosomiasis, polyP is synthesized by the vacuolar transporter chaperone (VTC) complex [2], which is located in the acidocalcisome [3], an acidic organelle that stores calcium and other cations together with Pi, inorganic pyrophosphate (PPi), and polyP [4]. A third endopolyphosphatase that has been described in yeast is diadenosine and diphosphoinositol polyphosphate phosphohydrolase (Ddp1) (YOR163W) [13], initially described as diadenosine hexaphosphate and diphosphoinositol polyphosphate hydrolase (DIPP) [14,15]. Ddp is localized in the cytosol and nucleus [16] and belongs to the Nudix (nucleoside diphosphate-linked moiety X) hydrolase family, which is characterized by a MuT motif or Nudix box of 23 amino acids (GX5EX7REUXEEXGU) where U is a bulky aliphatic residue and X is any amino acid [17]. Ddp and its human homologs DIPPs, License 4.0 (CC BY)
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