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Kinetic Regulation of Adenylate Kinases from Muscle, Liver, and Hepatoma¹

Department of Obstetrics and Gynecology, University of Florida College of Medicine, Gainesville, Florida 32610 [T. K. P., W.E.C.], and Department of Biochemistry, Howard University College of Medicine, Washington, D. C. 20001 [H. P. M.]

Homogeneous preparations of the major adenylate kinases (EC 2.7.4.3) from rat liver, skeletal muscle, and Morris hepatoma 3924A have been compared kinetically. At concentrations of 50 mM, most citric acid-cycle intermediates activated the enzyme from each tissue. The activation constants of citrate for liver, muscle, and hepatoma adenylate kinases were 0.09, 0.26, and 15.8 mM, respectively. It is likely that citrate could effect the in vivo functioning of liver adenylate kinase.

Apparent Michaelis constants were 1.9, 6.2, and 18.0 mM for adenosine 5'-monophosphate and were 7.0, 10.0, and 33.0 mM for adenosine 5'-triphosphate (ATP) with liver, muscle, and hepatoma enzymes, respectively. Citrate decreased the K_m (ATP) with liver and muscle adenylate kinases; it decreased the K_m (adenosine 5'-monophosphate) with only the muscle adenylate kinase.

Most Hill plot-slope values (for ATP) were near 1.5, indicating partial ligand-ligand cooperativity. Only in the presence of citrate, and with ATP as the variable substrate, was the slope value greater than two for liver adenylate kinase.

Several kinetic studies, in which modulation of the purified enzymes by nucleoside 5'-diphosphates, free fatty acids, mercurial reagents, and detergent, indicate that all three adenylate kinases have distinct and unique kinetic properties.

¹ This research was supported by Grants CA-11818, CA-10729, and CA-10906 from the NIH, and Grant F73-UF4 from the Florida Division of the American Cancer Society.

² Recipient of NIH Research Career Development Award CA-70187.

Received 4/15/74. Accepted 8/ 2/74.