Abstract
The pyruvate kinase of Trypanosoma brucei can be purified to homogeneity in one step by affinity elution from a phosphocellulose column with the substrate phosphoenolpyruvate (PEP) and the allosteric activator fructose-2,6-diphosphate (FDP). The purified enzyme has a specific activity of 175 μmol min-1 (mg protein)-1 and a subunit molecular mass of 59 kDa as judged by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Kinetic studies of the pure enzyme show that an increase in the PEP concentration decreases the apparent Km for adenosine diphosphate (ADP) and that an increase in the ADP concentration decreases the half saturation point (S0.5) for PEP. Likewise, the allosteric activator FDP decreases both the apparent Km for ADP and the S0.5 for PEP. ADP concentrations above 0.2 mM inhibit trypanosomal pyruvate kinase.
Original language | English (US) |
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Pages (from-to) | 141-147 |
Number of pages | 7 |
Journal | Molecular and Biochemical Parasitology |
Volume | 31 |
Issue number | 2 |
DOIs | |
State | Published - Nov 1988 |
Externally published | Yes |
Keywords
- Affinity elution
- Bioenergetics
- Enzyme kinetics
- Enzyme purification
- Pyruvate kinase
- Trypanosoma brucei
ASJC Scopus subject areas
- Parasitology
- Molecular Biology