Abstract

Translation elongation factor EF1A delivers aminoacyl-tRNA to the ribosome in a GTP-bound form, and is released from the ribosome in a GDP-bound form. This association/dissociation cycle proceeds efficiently via a marked conformational change in EF1A. EF1A function is dependent on the ribosomal “stalk” protein of the ribosomal large subunit, although the precise mechanism of action of the stalk on EF1A remains unclear. Here, we clarify the binding mode of archaeal stalk aP1 to GTP-bound aEF1A associated with aPelota. Intriguingly, the C-terminal domain (CTD) of aP1 binds to aEF1A•GTP with a similar affinity to aEF1A•GDP. We have also determined the crystal structure of the aP1-CTD•aEF1A•GTP•aPelota complex at 3.0 Å resolution. The structure shows that aP1-CTD binds to a space between domains 1 and 3 of aEF1A. Biochemical analyses show that this binding is crucial for protein synthesis. Comparison of the structures of aP1-CTD•aEF1A•GTP and aP1-CTD•aEF1A•GDP demonstrates that the binding mode of aP1 changes markedly upon a conformational switch between the GTP- and GDP-bound forms of aEF1A. Taking into account biochemical data, we infer that aP1 employs its structural flexibility to bind to aEF1A before and after GTP hydrolysis for efficient protein synthesis.

Highlights

  • Protein synthesis on the ribosome is regulated by the action of several translational GTPase factors on the ribosome with their action being coupled to GTP hydrolysis[1,2,3]

  • The results suggest that, despite the large conformational change of aEF1A before and after GTP hydrolysis, interaction between aP1-C-terminal domain (CTD) and aEF1A is maintained by switching of binding modes

  • The ribosomal stalk protein plays an important role in the recruitment of the GTP-bound form of elongation factor EF-Tu in bacteria, eEF1A in eukaryotes, and aEF1A in archaea, and in the delivery of the associated aminoacyl-tRNA to the A site of the ribosome

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Summary

Introduction

Protein synthesis on the ribosome is regulated by the action of several translational GTPase factors on the ribosome with their action being coupled to GTP hydrolysis[1,2,3]. The archaeal GTPase factor aEF1A (eEF1A in eukaryotes, EF-Tu in bacteria) delivers aminoacyl-tRNA to the ribosome in its GTP-bound form[4]. After codon recognition by the aminoacyl-tRNA and GTP hydrolysis, aEF1A is released from the ribosome in its GDP-bound form. The progress of the ribosomeaEF1A association/dissociation cycles mediated by GTP hydrolysis is related to efficient and accurate translation[3]. Domains 1 and 3, to which aP1-CTD binds in aP1aEF1AGDP (open conformation), is completely disrupted. The stalk is expected to bind to aEF1AGTP in a completely different mode to that when it is bound to aEF1AGDP. No data is available on the structure of the aP1aEF1AGTP complex

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