S-adenosylmethionine: Protein-arginine N-methyltransferase from bovine fetal liver - Purification and characterization

S-Adenosyl-L-methionine:protein arginine N-methyltransferase (Protein methylase I, PM-I)was purified from bovine fetal liver approximately 1,600-fold with an yield of 4%. The partially purified enzyme preparation showed 4 bands on SDS-PAGE. The enzyme has an optimum pH at around 7.8, and the activity was completely lost on heating at 50 degrees C for 5 min. Among substrate proteins used in vitro assay, histone type II-AS and 16 kDa endogenous protein in the isolated liver nuclei served as excellent substrates. The ratio of N-G-methylarginines produced by the enzyme with either histone type II-AS or isolated nuclei were different; N-G,N-G-dimethylarginine was the major methylated arginine with nuclei as a substrate, whereas N-G-MMA was the major product with histone type II-AS as a substrate. Cu2+ ion was the most potent inhibitor among various divalent cations tested, and 60% of the enzyme activity was inhibited at 50 mM concentration of Cu2+ (Ki = 4X10(-5) M). The Km value for AdoMet was 4X10(-6) M and Ki value for sinefungin was 8x10(-5) M. PM-I preparations at various stages of the enzyme purification invariably methylated 16 kDa protein in the nuclei used as in vitro substrate. This consistency of 16 kDa protein methylation during the enzyme purification suggests that a single enzyme is involved in the methylation of 16 kDa protein and that there might exist a 16 kDa protein specific PM-I.