Cooperative effects in the binding of pyridoxal 5'-phosphate to mitochondrial apo-aspartate aminotransferase.
Titrations of mitochondrial apo-aspartate aminotransferase with pyridoxal 5'-phosphate in the presence of AMP, contrary to what has been observed in the case of the cytosolic isoenzyme [(1983) FEBS Lett. 153, 98-102], show sigmoidal isotherms, with Hill coefficients ranging from nH = 1.4, in the absence of AMP, to nH = 1.8, in the presence of 5.9 mM AMP. The experimental data were successfully fitted by the Monod-Wyman- Changeaux model. The best fit, in the absence of AMP, was obtained with L = 30, KR = 4.72 X 10(-7) M and KT = 1.18 X 10(-5) M. Binding curves in the presence of AMP fit the model by keeping KR as a constant. This implies that AMP could bind to the apoenzyme only in the T state. In contrast, binding curves in the presence of phosphate ion (Pi) showed a less pronounced cooperativity, the Hill coefficient dropping to nH = 1.0 in the presence of 0.1 mM Pi. The above results suggest a regulatory role of AMP and Pi in the reconstitution of aspartate aminotransferase.
D000249:Adenosine Monophosphate / Q000378:metabolism / Q000494:pharmacology; D000818:Animals; D001051:Apoenzymes / Q000378:metabolism*; D001059:Apoproteins / Q000378:metabolism*; D001219:Aspartate Aminotransferases / Q000378:metabolism*; D003600:Cytosol / Q000201:enzymology; D046911:Macromolecular Substances; D008929:Mitochondria, Heart / Q000201:enzymology*; D010710:Phosphates / Q000494:pharmacology; D011485:Protein Binding; D011487:Protein Conformation / Q000187:drug effects; D011732:Pyridoxal Phosphate / Q000378:metabolism*; D013552:Swine
D001051:Apoenzymes; D001059:Apoproteins; D046911:Macromolecular Substances; D010710:Phosphates; D000249:Adenosine Monophosphate; D011732:Pyridoxal Phosphate; D001219:Aspartate Aminotransferases