Coenzyme reorientations in the active sites of aspartate aminotransferase isoenzymes studied by linear dichroism method.
Cytosolic and mitochondrial pig heart aspartate aminotransferases (cAspAT and mAspAT) and chicken heart cAspAT have been oriented in a compressed slab of polyacrylamide gel and their linear dichroism LD spectra have been recorded. The coenzyme's tilt angles in the active sites of chicken cAspAT and pig mAspAT and their quasisubstrate complexes imitating catalytic intermediates have been computed. The computations are based on reduced linear dichroism values (delta A/A), the known directions of the transition dipole moments in the coenzyme ring and atomic coordinates of the coenzyme obtained by X-ray crystallography. It has been found that formation of the enzyme complex with glutarate and protonation of the internal pyridoxal-lysine aldimine induce reorientations of the coenzyme. As a result of protonation, the coenzyme ring tilts by 27 degrees in cAspAT and 13 degrees in mAspAT. Formation of the external aldimine with 2-methylaspartate is accompanied by tilting of the coenzyme ring by 44 degrees in cAspAT and 39 degrees in mAspAT. For the quinonoid complex with erythro-3-hydroxyaspartate, the tilt angles were found to be 63 degrees in cAspAT and 53 degrees in mAspAT. It is inferred that the basic features of the active site dynamics are similar in the three AspAT's studied. The differences in the coenzyme tilt angles between cAspAT and mAspAT may be linked to catalytic and structural peculiarities of the isoenzymes.
Rosenberg
M V
MV
Institute of Molecular Biology, USSR Academy of Sciences, Moscow.
Makarov
V L
VL
Torchinsky
Y M
YM
D000818:Animals; D001219:Aspartate Aminotransferases; D001665:Binding Sites; D002645:Chickens; D003600:Cytosol / Q000201:enzymology; D007527:Isoenzymes; D008929:Mitochondria, Heart / Q000201:enzymology; D011487:Protein Conformation; D011732:Pyridoxal Phosphate; D013057:Spectrum Analysis; D013552:Swine
D007527:Isoenzymes; D011732:Pyridoxal Phosphate; D001219:Aspartate Aminotransferases