Specific labeling of cytosolic and mitochondrial aspartate aminotransferases.
The apoisozymes of cytosolic and mitochondrial aspartate aminotransferase are both irreversibly inhibited by alpha-N-fluorodinitrophenyl-beta-N-phosphopyridoxyldiaminopropi onate, an affinity-labeling reagent analog of the coenzyme. Analysis of the modified peptides shows that the active-site Lys-258, which in the holoenzyme binds the coenzyme pyridoxal 5'-phosphate, is labeled in both isozymes. Comparison with the results obtained using the parent compound 4'-N-fluorodinitrophenylpyridoxamine 5'-phosphate, which labels only the cytosolic enzyme, provides information about differences in active-site reactivity and geometry. Labeling external to the active site occurs in both isozymes. In the cytosolic enzyme the very reactive Cys-45 is modified, in the mitochondrial enzyme the surface residue Lys-342 reveals a peculiar reactivity.
D000345:Affinity Labels / Q000138:chemical synthesis*; D000818:Animals; D001219:Aspartate Aminotransferases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D055598:Chemical Phenomena; D002621:Chemistry; D003600:Cytosol / Q000201:enzymology*; D004139:Dinitrofluorobenzene / Q000031:analogs & derivatives / Q000138:chemical synthesis; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000378:metabolism*; D008929:Mitochondria, Heart / Q000201:enzymology*; D009578:Nitrobenzenes; D010446:Peptide Fragments / Q000302:isolation & purification; D013552:Swine
D000345:Affinity Labels; D007527:Isoenzymes; D009578:Nitrobenzenes; D010446:Peptide Fragments; C044341:alpha-N-fluorodinitrophenyl-beta-N-phosphopyridoxyldiaminopropionate; D004139:Dinitrofluorobenzene; D001219:Aspartate Aminotransferases