Enzymatic characterization of a class II lysyl-tRNA synthetase, LysS, from Myxococcus xanthus

Lysyl-tRNA synthetases efficiently produce diadenosine tetraphosphate (Ap₄A) from lysyl-AMP with ATP in the absence of tRNA. We characterized recombinant class II lysyl-tRNA synthetase (LysS) from Myxococcus xanthus and found that it is monomeric and requires Mn²⁺ for the synthesis of Ap₄A. Surprisingly, Zn²⁺ inhibited enzyme activity in the presence of Mn²⁺. When incubated with ATP, Mn²⁺, lysine, and inorganic pyrophosphatase, LysS first produced Ap₄A and ADP, then converted Ap₄A to diadenosine triphosphate (Ap₃A), and finally converted Ap₃A to ADP, the end product of the reaction. Recombinant LysS retained Ap₄A synthase activity without lysine addition. Additionally, when incubated with Ap₄A (minus pyrophosphatase), LysS converted Ap₄A mainly ATP and AMP, or ADP in the presence or absence of lysine, respectively. These results demonstrate that M. xanthus LysS has different enzymatic properties from class II lysyl-tRNA synthetases previously reported.