Table 1.
Effects of 0.1 mM SP, GP and AP on the activities of the enzymes of central metabolism, transforming 2-oxo acids and/or their analogues.
| Enzyme | Substrate concentration | Source | Specific activities | |||
|---|---|---|---|---|---|---|
| Ctrl | 0.1 mM SP | 0.1 mM GP | 0.1 mM AP | |||
| OADHC/OGDHC | 0.01 mM 2-oxoadipate | Enzyme-enriched fraction from rat liver | 0.96 ± 0.10 | 0.59 ± 0.04 | 0 | 0.09 ± 0.01 |
| 0.2 mM 2-oxoglutarate | 14.9 ± 0.4 | 2.5 ± 0.2 | 0.2 ± 0.2 | 12.1 ± 0.5 | ||
| 0.01 mM 2-oxoadipate | Enzyme-enriched fraction from rat heart | 1.23 ± 0.04 | 0 | 0 | 0.87 ± 0.36 | |
| 0.2 mM 2-oxoglutarate | 123 ± 1 | 50 ± 3 | 52 ± 1 | 119 ± 2 | ||
| PDHC | 0.2 mM pyruvate | Enzyme-enriched fraction from rat heart | 163 ± 4 | 148 ± 2 | 144 ± 1 | 156 ± 7 |
| BCDHC | 0.2 mM 3-methyl-2-oxovalerate | Enzyme-enriched fraction from rat liver | 1.81 ± 0.05 | 1.70 ± 0.06 | 1.75 ± 0.04 | 1.93 ± 0.11 |
| GDH | 0.2 mM 2-oxoglutarate | Purified from bovine liver | 81.9 ± 1.3 | 74.8 ± 0.5 | 69.3 ± 0.8 | 73.7 ± 3.1 |
| Rat liver homogenate | 27.3 ± 0.4 | 28.4 ± 0.3 | 28.3 ± 0.0 | 26.3 ± 0.4 | ||
| MDH | 0.2 mM oxaloacetate | Rat liver homogenate | 345 ± 11 | 315 ± 22 | 369 ± 14 | 379 ± 11 |
| Rat heart homogenate | 471 ± 5 | 500 ± 8 | 518 ± 7 | 517 ± 7 | ||
| LDH | 0.2 mM lactate | Purified from rabbit muscle | 219 ± 1 | 217 ± 2 | 215 ± 8 | 211 ± 5 |
| PK | 0.2 mM phospho-enolpyruvate | Purified from rabbit muscle | 27.3 ± 1.0 | 24.1 ± 0.3 | 25.6 ± 0.8 | 26.9 ± 1.4 |
| GOT | 2 mM 2-oxoglutarate | Rat liver homogenate | 16.7 ± 0.7 | 19.6 ± 0.2 | 20.3 ± 0.2 | 21.4 ± 0.4 |
| Rat heart homogenate | 30.4 ± 0.6 | 31.7 ± 1.2 | 28.7 ± 1.0 | 30.9 ± 0.3 | ||
| GPT | Rat liver homogenate | 1.83 ± 0.06 | 1.74 ± 0.41 | 1.49 ± 0.23 | 1.91 ± 0.10 | |
| NADP+-IDH | 2 mM isocitrate | Rat liver homogenate | 5.30 ± 0.11 | 4.31 ± 0.06 | 5.13 ± 0.03 | 5.36 ± 0.03 |
| Rat heart homogenate | 1.20 ± 0.02 | 1.00 ± 0.01 | 1.29 ± 0.11 | 1.28 ± 0.04 | ||
| NADP+-ME | 2 mM malate | Rat liver homogenate | 0.085 ± 0.002 | 0.067 ± 0.002 | 0.078 ± 0.005 | 0.102 ± 0.002 |
All enzyme assays were performed under standard assay conditions (see Material and Methods) at the indicated concentrations of the 2-oxo substrates or their analogues. Specific activities of the enzymes are given as mean ± SEM in nmol/min per mg of protein for the enzyme-enriched fractions, in µmol/min per mg of protein for purified enzymes, or in µmol/min per g of tissue fresh weight (gFW) for homogenates. The following abbreviations are used: OADHC – 2-oxoadipate dehydrogenase complex, OGDHC – 2-oxoglutarate dehydrogenase complex, PDHC – pyruvate dehydrogenase complex, BCDHC – branched-chain 2-oxo acid dehydrogenase complex, GDH – glutamate dehydrogenase, MDH – malate dehydrogenase, LDH – lactate dehydrogenase, PK – pyruvate kinase, GOT – glutamate oxaloacetate transaminase, GPT – glutamate pyruvate transaminase, NADP+-IDH – NADP+-dependent isocitrate dehydrogenase, NADP+-ME – NADP+-dependent malic enzyme.