Interaction of mitochondrial elongation factor Tu with aminoacyl-tRNA and elongation factor Ts

Elongation factor (EF) Tu promotes the binding of aminoacyl-tRNA (aa-tRNA) to the acceptor site of the ribosome, This process requires the formation of a ternary complex (EF-Tu.GTP.aa-tRNA). EF-Tu is released from the ribosome as an EF-Tu GDP complex. Exchange of GDP for GTP is carried out through the formation of a complex with EF-Ts (EF-Tu.Ts). Mammalian mitochondrial EF-Tu (EF-Tu(mt)) differs from the corresponding prokaryotic factors in having a much lower affinity for guanine nucleotides, To further understand the EF-Tu(mt) subcycle, the dissociation constants for the release of aa-tRNA from the ternary complex (K-tRNA) and for the dissociation of the EF-Tu.Ts-mt complex (K-Ts) were investigated. The equilibrium dissociation constant for the ternary complex was 18 +/- 4 nM, which is close to that observed in the prokaryotic system. The kinetic dissociation rate constant for the ternary complex was 7.3 x 10(-4) s(-1), which is essentially equivalent to that observed for the ternary complex in Escherichia coli. The binding of EF-Tu(mt) to EF-Ts-mt is mutually exclusive with the formation of the ternary complex, K-Ts was determined by quantifying the effects of increasing concentrations of EF-Ts-mt on the amount of ternary complex formed with EF-Tu(mt). The value obtained for K-Ts (5.5 +/- 1.3 nM) is comparable to the value K-tRNA.