Characterization of a homogeneous complex of arginyl- and lysyl-tRNA synthetase: zinc and adenosine 5'-phosphate dependent synthesis of diadenosine 5',5'''-P1,P4-tetraphosphate

Abstract

Zinc, adenosine 5'-phosphate (AMP), and pyrophosphatase greatly stimulate the synthesis of diadenosine 5',5'''-P1,P4-tetraphosphate (Ap4A) by rat liver lysyl-tRNA synthetase. The synthesis of Ap4A does not require lysine; thus the lysyl-adenylate complex is not required. The substrates have been determined to be adenosine 5'-triphosphate (ATP) and AMP with apparent Km values of 2.1 mM and 1.5 mM, respectively. A zinc-dependent hydrolysis of ATP and AMP has been shown to be associated with the synthetase. In the presence of zinc there is a direct correlation between both the amount of AMP formed and the amount of Ap4A synthesized by lysyl-tRNA synthetase. Ap4A acts as a competitive inhibitor for ATP in the aminoacylation reaction of lysyl-tRNA synthetase with a KI of 2.5 microM. Concentrations of Ap4A up to 12.5 microM do not inhibit the synthesis of Ap4A by lysyl-tRNA synthetase. This suggests that there may be more than one binding site for ATP on the enzyme.