Details

Autor(en) / Beteiligte

• Hammond, D.J.
• Bowman, I.B.R.

Titel

Studies on glycerol kinase and its role in ATP synthesis in Trypanosoma brucei

Ist Teil von

• Molecular and biochemical parasitology, 1980-01, Vol.2 (2), p.77-91

Ort / Verlag

Netherlands: Elsevier B.V

Erscheinungsjahr

1980

Quelle

Elsevier Journal Backfiles on ScienceDirect (DFG Nationallizenzen)

Beschreibungen/Notizen

• Glycerol kinase of Trypanosoma brucei has been shown to be capable of catalysing sn-glycerol-3-phosphate dependent ADP phosphorylation for ATP generation. The rate of this reaction ( V r) is sufficient to account for the observed rate of glycerol production from anaerobic glucose metabolism by intact cells and to account for net ATP synthesis. Glycerol kinase has been purified by preparing a post-nuclear, particulate fraction and solubilizing the enzyme with 0.5% (w/v) Triton X-100. This treatment results in a 3.5-fold increase in total activity, demonstrating the latent nature of particulate glycerol kinase, and an overall 10-fold increase in specific activity in the soluble fraction. The ratio of the velocities of the forward ( V f) to reverse ( V r) reactions of this enzyme is altered from 21 to 170 upon solubilization. The Michaelis constants for the solubilized enzyme are K m ADP = 0.12 ± 0.04 mM, K m G-3- P = 5.12 ± 1.47 mM, K m glycerol = 0.12 ± 0.05 and K m ATP = 0.19 ± 0.04 mM. Endogenous hexokinase acts as an ATP trap favouring ATP synthesis from sn-glycerol-3-phosphate and ADP. This can be demonstrated in reconstituted systems using trypanosome glycerol kinase and varying hexokinase activities. Mass action inhibition of ATP synthesis by glycerol is more marked with lower hexokinase activities. High glycerol kinase activity (> 0.5 μmol/min/mg protein) has been found in the T. brucei complex of trypanosomes that produce glycerol anaerobically whereas only low activities (⩽ 0.03 μmol/min/mg protein) are present in Trypanosoma cruzi, Trypanosoma lewisi and Crithidia fasciculata, organisms that do not produce glycerol. Trypanosoma congolense has a glycerol kinase activity of 0.17 μmol/min/mg protein and shows poorer ATP synthesis from anaerobic glucose metabolism than organisms of the T. brucei complex.