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Cooperative effects in the binding of pyridoxal 5′‐phosphate to mitochondrial apo‐aspartate aminotransferase
Author(s) -
Garzillo A.M.,
Marino G.,
Pispisa B.
Publication year - 1984
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/0014-5793(84)81317-4
Subject(s) - pyridoxal 5 phosphate , cooperativity , pyridoxal , cooperative binding , chemistry , pyridoxal phosphate , pi , phosphate , cytosol , biochemistry , enzyme , stereochemistry , cofactor
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 n H = 1.4, in the absence of AMP, to n H = 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, K R = 4.72 × 10 −7 M and K T = 1.18 × 10 −5 M. Binding curves in the presence of AMP fit the model by keeping K R 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 (P i ) showed a less pronounced cooperativity, the Hill coefficient dropping to n H = 1.0 in the presence of 0.1 mM P i . The above results suggest a regulatory role of AMP and P i in the reconstitution of aspartate aminotransferase.