Roles of triosephosphate isomerase and aerobic metabolism in Trypanosoma brucei

Abstract

Kinetoplastid protozoa compartmentalize the first seven enzymes of glycolysis and two enzymes of glycerol metabolism in a microbody, the glycosome. While in its mammalian host, Trypanosoma brucei depends entirely on glucose for ATP generation. Under aerobic conditions, most of the glucose is metabolized to pyruvate. Aerobic metabolism depends on the activities of glycosomal triosephosphate isomerase and a mitochondrial glycerophosphate oxidase, and on glycerophosphate ↔ dihydroxyacetone phosphate exchange across the glycosomal membrane. Using a combination of genetics and computer modelling, we show that triosephosphate isomerase is probably essential for bloodstream trypanosome survival, but not for the insect-dwelling procyclics, which preferentially use amino acids as an energy source. When the enzyme level decreased to about 15% of that of the wild-type, the growth rate was halved. Below this level, a lethal rise in dihydroxyacetone phosphate was predicted. Expression of cytosolic triosephosphate isomerase inhibited cell growth. Attempts to knockout the trypanosome alternative oxidase genes (which are needed for glycerophosphate oxidase activity) were unsuccessful, but when we lowered the level of the corresponding mRNA by expressing a homologous double-stranded RNA, oxygen consumption was reduced fourfold and the rate of trypanosome growth was halved.

A proteomic approach to identify developmentally regulated proteins in Leishmania infantum

“…In promastigotes, these values were at 0.520 mM and at 0.328 U/mg, respectively ( Table 3). The K m value of the L. infantum TIM in promastigotes was in close agreement to that reported for the L. mexicana (0.3 mM) [50] and Trypanosoma brucei (0.25 mM) [57] homologs. The kinetic properties of TIM in Leishmania amastigotes are reported here for the first time.…”

“…While in its mammalian host, Leishmania depends mainly on glucose and on fatty acids for generating ATP. It has been shown recently that TIM is probably essential for bloodstream trypanosome survival but not for procyclics, which preferentially use amino acids as an energy source [57]. Indeed, as TIM decreased below 15% of wild-type activity, all fluxes (pyruvate, glycerol and glucose) were not measurable [57].…”

“…It has been shown recently that TIM is probably essential for bloodstream trypanosome survival but not for procyclics, which preferentially use amino acids as an energy source [57]. Indeed, as TIM decreased below 15% of wild-type activity, all fluxes (pyruvate, glycerol and glucose) were not measurable [57]. We showed that in L. infantum amastigotes, the specific activity of TIM was two-fold higher compared to the promastigote form and this independently of the substrate concentration ( Table 3).…”

Triosephosphate isomerase: a highly evolved biocatalyst

“…In the human genome, there is only one gene encoding for TIM [36]. In most organisms, TIM is located in the cytosol, but in trypanosomes and leishmania, it is found both in the cytosol as well as in the glycosome (which is a peroxisome-like microbody, important for the glycolysis in these organisms) [37]. Also in these organisms only one gene is found [38].…”

Intermediary Metabolism in Protists: a Sequence-based View of Facultative Anaerobic Metabolism in Evolutionarily Diverse Eukaryotes

“…the asexual blood stages of malarial parasites; Sherman 1979) or is itself excreted from cells (e.g. the African trypanosome Trypanosoma brucei , which uses a dihydroxyacetone-glycerol-3-phosphate shuttle to support NADH oxidation; Brohn and Clarkson 1978; Helfert et al 2001; Hellemond et al 2005). This emphasis of glycolysis as an energy source in some protists is reminiscent of the situation often seen in proliferating mammalian cells, including tumour cells, where aerobic glycolysis and the production of lactate is used to sustain cell growth (‘the Warburg effect’); here, the preference for aerobic glycolysis over oxidative phosphorylation to sustain ATP production may even be a pre-requisite for sustaining flux through the biosynthetic pathways necessary to sustain rapid cell proliferation (Vander Heiden et al 2009).…”