Adenosine 3′,5′-Monophosphate-dependent Protein Kinase from Bovine Anterior Pituitary Gland: I. PROPERTIES

Abstract

Abstract Adenosine 3',5'-monophosphate-dependent protein kinase was purified from bovine anterior pituitary gland by pH and ammonium sulfate precipitations and chromatography on sulfoethyl-Sephadex. Maximal rates of histone phosphorylation were observed in the presence and absence of adenosine 3',5'-monophosphate (cyclic AMP) at pH 6.5 and 7.0, respectively. The apparent Km value for histone was 125 µg per ml in the presence and absence of cyclic AMP. The apparent Km values for ATP in the presence and absence of cyclic AMP were 1.7 and 1.3 x 10-5 m, respectively. Half-maximal stimulation by the cyclic nucleotide was observed at 2.5 x 10-8 m. Maximal activation of protein kinase activity was obtained at much higher concentrations of the other cyclic nucleotides. No significant phosphorylation of histones occurred when γ-labeled UTP, CTP, or GTP was used instead of ATP. The enzyme requires both Mg2+ and ATP-Mg2- for activity. Magnesium decreases the apparent Km of the enzyme for ATP and increases the Vmax of enzymatic activity. The two peaks of cyclic AMP-dependent protein kinase activity observed on sucrose gradients are converted to a single light peak in the presence of cyclic AMP.