Review: Translational GTPases

Abstract

Translational GTPases (trGTPases) play key roles in facilitating protein synthesis on the ribosome. Despite the high degree of evolutionary conservation in the sequences of their GTP‐binding domains, the rates of GTP hydrolysis and nucleotide exchange vary broadly between different trGTPases. EF‐Tu, one of the best‐characterized model G proteins, evolved an exceptionally rapid and tightly regulated GTPase activity, which ensures rapid and accurate incorporation of amino acids into the nascent chain. Other trGTPases instead use the energy of GTP hydrolysis to promote movement or to ensure the forward commitment of translation reactions. Recent data suggest the GTPase mechanism of EF‐Tu and provide an insight in the catalysis of GTP hydrolysis by its unusual activator, the ribosome. Here we summarize these advances in understanding the functional cycle and the regulation of trGTPases, stimulated by the elucidation of their structures on the ribosome and the progress in dissecting the reaction mechanism of GTPases. © 2016 Wiley Periodicals, Inc. Biopolymers 105: 463–475, 2016.

Two proofreading steps amplify the accuracy of genetic code translation

“…All tRNAs (WT and mutants) were unmodified and based on native E. coli (A) tRNA Glu or (B) tRNA Phe (black with purple anticodon; tRNA modifications are in green) with changes in blue. EF-Tu·GDP·Pi from the ribosome-bound aa-tRNA but Pi release and conformational change of the factor (16). Therefore, such a proofreading step would not be expected to display the linear dependence of the proofreading factor F on the affinity parameter K A as predicted by our model (Fig.…”

“…1). By this mechanism, GTP hydrolysis on ribosome-bound EF-Tu first leads to a ribosome complex with aa-tRNA, EF-Tu·GDP, and inorganic phosphate, Pi, in which aa-tRNA is strongly bound to EF-Tu (16). After rapid release of Pi, EF-Tu changes conformation from the GTP to the GDP form.…”

“…Could, then, an alternative model with dissociation of aa-tRNA in complex with EF-Tu·GDP·Pi as discard reaction account for the present experimental results? According to this model, aa-tRNA would be discarded in complex with EF-Tu·GDP·Pi after GTP hydrolysis but before Pi release and conformational change of EF-Tu (16). Due to high affinity of aa-tRNA to EF-Tu before the conformational change of the factor, the forward rate constant in such a step would not be dissociation of A B Table S1.…”

“…Soon thereafter, GTP-driven proofreading of codon reading by aa-tRNAs on the mRNA-translating ribosome was discovered (8,9) and subsequently subjected to extensive research (10)(11)(12)(13). Although it was early recognized that multistep proofreading confers higher accuracy and kinetic efficiency to substrate-selective, enzyme-catalyzed reactions than single-step proofreading (5,14,15), it has been taken for granted that there is but a single proofreading step in tRNA selection by the translating ribosome (16). Here, we present data showing that the proofreading factor (F), by which the accuracy (A) is amplified from its initial selection value (I) by aa-tRNA in ternary complex with EF-Tu and GTP, increases linearly with increasing association equilibrium constant, K A , for aa-tRNA binding to EFTu.…”

Common mechanisms of catalysis in small and heterotrimeric GTPases and their respective GAPs

“…All GTPases share a G-domain with five α-helices and a six-stranded β-sheet. They form the two large families described above, the heterotrimeric GTPases (Sprang, 2016) and the small GTPases (Mishra and Lambright, 2016), as well as the dynamin superfamily (Daumke and Praefcke, 2016) and translational GTPases (Maracci and Rodnina, 2016). The small GTPases are grouped into five subfamilies: Ras GTPases are mainly involved in cell growth (Cox and Der, 2010), Rho GTPases function in the regulation of the cytoskeleton (Sit and Manser, 2011), Rab (Stenmark, 2009) and Arf (Khan and Ménétrey, 2013) regulate vesicular transport, and Ran (Jamali et al, 2011) controls nuclear transport.…”