biostudies-literatureFernandez PLNational Institute of General Medical SciencesNIGMS NIH HHS2672-2676https://www.ebi.ac.uk/biostudies/studies/S-EPMC8436233biostudies-literatureUnknown60(35)The binding of adenosine 5'-triphosphate (ATP) and adenosine 5'-monophosphate (AMP) to adenylate kinase (AdK) drives closure of lids over the substrate adenosyl groups. We test the hypothesis that this conformational change activates AdK for catalysis. The rate constants for <i>Homo sapiens</i> adenylate kinase 1 (<i>Hs</i>AdK1)-catalyzed phosphoryl group transfer to AMP, <i>k</i>_cat/<i>K</i>_m = 7.0 × 10⁶ M^-1 s^-1, and phosphite dianion, (<i>k</i>_HPi)_obs ≤1 × 10^-4 M^-1 s^-1, show that the binding energy of the adenosyl group effects a ≥7.0 × 10¹⁰-fold rate acceleration of phosphoryl transfer from ATP. The third-order rate constant of <i>k</i>_cat/<i>K</i>_HPi<i>K</i>_EA = 260 M^-2 s^-1 for 1-(β-d-erythrofuranosyl)adenine (EA)-activated phosphoryl transfer to phosphite dianion was determined, and the isohypophosphate reaction product characterized by ³¹P NMR. The results demonstrate the following: (i) a ≥14.7 kcal/mol stabilization of the transition state for phosphoryl transfer by the adenosyl group of AMP and a ≥2.6 × 10⁶-fold rate acceleration from the EA-driven conformational change and (ii) the recovery of ≥8.7 kcal/mol of this transition state stabilization for EA-activated phosphoryl transfer from ATP to phosphite.BiochemistryAdenylate Kinase-Catalyzed Reaction of AMP in Pieces: Enzyme Activation for Phosphoryl Transfer to Phosphite Dianion.PMC8436233R35 GM134881GM134881R01 GM116921Richard JPFernandez PLfalseAdenylate Kinase-Catalyzed Reaction of AMP in Pieces: Enzyme Activation for Phosphoryl Transfer to Phosphite Dianion.The binding of adenosine 5'-triphosphate (ATP) and adenosine 5'-monophosphate (AMP) to adenylate kinase (AdK) drives closure of lids over the substrate adenosyl groups. We test the hypothesis that this conformational change activates AdK for catalysis. The rate constants for <i>Homo sapiens</i> adenylate kinase 1 (<i>Hs</i>AdK1)-catalyzed phosphoryl group transfer to AMP, <i>k</i>_cat/<i>K</i>_m = 7.0 × 10⁶ M^-1 s^-1, and phosphite dianion, (<i>k</i>_HPi)_obs ≤1 × 10^-4 M^-1 s^-1, show that the binding energy of the adenosyl group effects a ≥7.0 × 10¹⁰-fold rate acceleration of phosphoryl transfer from ATP. The third-order rate constant of <i>k</i>_cat/<i>K</i>_HPi<i>K</i>_EA = 260 M^-2 s^-1 for 1-(β-d-erythrofuranosyl)adenine (EA)-activated phosphoryl transfer to phosphite dianion was determined, and the isohypophosphate reaction product characterized by ³¹P NMR. The results demonstrate the following: (i) a ≥14.7 kcal/mol stabilization of the transition state for phosphoryl transfer by the adenosyl group of AMP and a ≥2.6 × 10⁶-fold rate acceleration from the EA-driven conformational change and (ii) the recovery of ≥8.7 kcal/mol of this transition state stabilization for EA-activated phosphoryl transfer from ATP to phosphite.2021-01-01T00:00:00Z2021 Sep2025-04-04T09:37:49.876Z2025-02-18T23:49:06.049ZS-EPMC84362333443577610.1021/acs.biochem.1c00535