CHARACTERIZATION OF THE ATP-SPECIFIC AND GTP-SPECIFIC SUCCINYL-COA SYNTHETASES IN PIGEON - THE ENZYMES INCORPORATE THE SAME ALPHA-SUBUNIT

Two succinyl-CoA synthetases, one highly specific for GTP/GDP and the other for ATP/ADP, have been purified to homogeneity from pigeon liver and breast muscle. The two enzymes are differentially distributed in pigeon, with only the GTP-specific enzyme detected in liver and the AT P-specific enzyme in breast muscle, Based on assays in the direction o f CoA formation, the ratios of GTP specific to ATP-specific activities in kidney, brain, and heart are similar to 7, 1, and 0.1, respectivel y. Both enzymes have the characteristic alpha- and beta-subunits found in other succinyl-CoA synthetases. Studies of the alpha-subunit by el ectrophoresis, mass spectrometry, reversed-phase high performance liqu id chromatography, and peptide mapping showed that it was the same in the two enzymes. Characterization of the beta-subunits by the same met hods indicated that they were different, with the tryptic peptide maps providing evidence that the beta-subunits likely differ along their e ntire sequences. Because the two succinyl-CoA synthetases incorporate the same alpha-subunit, the determinants of nucleotide specificity mus t reside within the beta-subunit, Determination of the apparent Michae lis constants showed that the affinity of the GTP-specific enzyme for GDP is greater than that of the ATP-specific enzyme for ADP (7 versus 250 mu M). Rather large differences in apparent K-m values were also o bserved for succinate and phosphate.