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-Tumt) differs from the corresponding prokaryotic factors in having a much lower affinity for guanine nucleotides. To further understand the EF-Tumt subcycle, the dissociation constants for the release of aa-tRNA from the ternary complex (KtRNA) and for the dissociation of the EF-Tu·Tsmt complex (KTs) 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 × 10−4 s−1, which is essentially equivalent to that observed for the ternary complex inEscherichia coli. The binding of EF-Tumt to EF-Tsmt is mutually exclusive with the formation of the ternary complex. KTs was determined by quantifying the effects of increasing concentrations of EF-Tsmt on the amount of ternary complex formed with EF-Tumt. The value obtained for KTs(5.5 ± 1.3 nm) is comparable to the value ofKtRNA.