Importance of individual amino acids in the Switch I region in eEF2 studied by functional complementation in S. cerevisiae

Bartish, Galyna; Nygård, Odd

doi:10.1016/j.biochi.2008.01.006

Cited by 6 publications

(4 citation statements)

References 66 publications

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“…Among five GTP binding motifs, Switch I always retained its flexibility, and P-loop and Switch II changed their conformation, responding to the binding-state, whereas the G4 and the G5 motifs exhibited similar conformations in the different forms. In this study, it was clearly shown that the α-phosphate of GDP was recognized not only by P-loop, but also by the side chain of the residue R71 of Switch I (Figure 2B ), which is conserved in all organisms, and involved in increasing the GTPase activity on the ribosome and promoting translocation ( 55 , 56 ).…”

Section: Discussionmentioning

confidence: 76%

The C-terminal helix of ribosomal P stalk recognizes a hydrophobic groove of elongation factor 2 in a novel fashion

Tanzawa

Kato

Girodat

et al. 2018

Nucleic Acids Research

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Archaea and eukaryotes have ribosomal P stalks composed of anchor protein P0 and aP1 homodimers (archaea) or P1•P2 heterodimers (eukaryotes). These P stalks recruit translational GTPases to the GTPase-associated center in ribosomes to provide energy during translation. The C-terminus of the P stalk is known to selectively recognize GTPases. Here we investigated the interaction between the P stalk and elongation factor 2 by determining the structures of Pyrococcus horikoshii EF-2 (PhoEF-2) in the Apo-form, GDP-form, GMPPCP-form (GTP-form), and GMPPCP-form bound with 11 C-terminal residues of P1 (P1C11). Helical structured P1C11 binds to a hydrophobic groove between domain G and subdomain G′ of PhoEF-2, where is completely different from that of aEF-1α in terms of both position and sequence, implying that such interaction characteristic may be requested by how GTPases perform their functions on the ribosome. Combining PhoEF-2 P1-binding assays with a structural comparison of current PhoEF-2 structures and molecular dynamics model of a P1C11-bound GDP form, the conformational changes of the P1C11-binding groove in each form suggest that in response to the translation process, the groove has three states: closed, open, and release for recruiting and releasing GTPases.

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Section: Discussionmentioning

confidence: 76%

The C-terminal helix of ribosomal P stalk recognizes a hydrophobic groove of elongation factor 2 in a novel fashion

Tanzawa

Kato

Girodat

et al. 2018

Nucleic Acids Research

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“…This mutant yeast cell grew extremely slowly, showed increased cell death and became temperature sensitive. Hence this ability to replace eEF2 in vivo showed that the structural rearrangement of Switch I necessary for eEF2 function which is similar in other eukaryote 17. Taylor and group recently provided a basis for explaining the stepwise mechanism of translocation by providing the structural data 19.…”

Section: Introductionmentioning

confidence: 93%

“…The eEF2 is a member of the G‐protein super family which undergoes conformational changes associated with binding of the guanosine nucleotide and hydrolysis of the bound GTP. These structural rearrangements affect the Switch I region of eEF2 which is also known as effector loop 17. Switch I region is a structural motif involved in coordinating the Mg 2+ ion found in the nucleotide‐binding pocket 18.…”

Section: Introductionmentioning

confidence: 99%

Eukaryotic elongation factor‐2 (eEF2): its regulation and peptide chain elongation

Kaul

Pattan

Rafeequi

2011

Cell Biochemistry & Function

181

119

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Regulation at the level of translation in eukaryotes is feasible because of the longer lifetime of eukaryotic mRNAs in the cell. The elongation stage of mRNA translation requires a substantial amount of energy and also eukaryotic elongation factors (eEFs). The important component of eEFs, i.e. eEF2 promotes the GTP-dependent translocation of the nascent protein chain from the A-site to the P-site of the ribosome. Mostly the eEF2 is regulated by phosphorylation and dephosphorylation by a specific kinase known as eEF2 kinase, which itself is up-regulated by various mechanisms in the eukaryotic cell. The activity of this kinase is dependent on calcium ions and calmodulin. Recently it has been shown that the activity of eEF2 kinase is regulated by MAP kinase signalling and mTOR signalling pathway. There are also various stimuli that control the peptide chain elongation in eukaryotic cell; some stimuli inhibit and some activate eEF2. These reports provide the mechanisms by which cells likely serve to slow down protein synthesis and conserve energy under nutrient deprived conditions via regulation of eEF2. The regulation via eEF2 has also been seen in mammary tissue of lactating cows, suggesting that eEF2 may be a limiting factor in milk protein synthesis. Regulation at this level provides the molecular understanding about the control of protein translocation reactions in eukaryotes, which is critical for numerous biological phenomenons. Further the elongation factors could be potential targets for regulation of protein synthesis like milk protein synthesis and hence probably its foreseeable application to synthetic biology.

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“…By contrast, EF-G promotes movement of mRNA and tRNA on the ribosome in steps that involve large-scale rearrangements of the ribosome (7)(8)(9)(10)(11). Biochemical and genetic experiments have shown that the GTPase centers of EF-Tu and EF-G, although highly conserved, are not interchangeable (12), whereas key amino acids in the GTPase active site of the eukaryotic translocase eEF2 can be mutated to those of EF-G and retain function (13). Furthermore, whereas EF-Tu hydrolyzes GTP rapidly only during accurate mRNA decoding, the GTPase activity of EF-G is greatly accelerated even by vacant ribosomes (14,15).…”

Section: Control Of Ribosomal Subunit Rotation By Elongation Factor Gmentioning

confidence: 99%

Control of Ribosomal Subunit Rotation by Elongation Factor G

Pulk

Cate

2013

Science

171

218

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Protein synthesis by the ribosome requires the translocation of transfer RNAs and messenger RNA by one codon after each peptide bond is formed, a reaction that requires ribosomal subunit rotation and is catalyzed by the guanosine triphosphatase (GTPase) elongation factor G (EF-G). We determined 3 Å resolution x-ray crystal structures of EF-G complexed with a non-hydrolyzable GTP analogue and bound to the Escherichia coli ribosome in different states of ribosomal subunit rotation. The structures reveal that EF-G binding to the ribosome stabilizes switch regions in the GTPase active site, resulting in a compact EF-G conformation that favors an intermediate state of ribosomal subunit rotation. These structures suggest that EF-G controls the translocation reaction by cycles of conformational rigidity and relaxation preceding and following GTP hydrolysis.

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Importance of individual amino acids in the Switch I region in eEF2 studied by functional complementation in S. cerevisiae

Cited by 6 publications

References 66 publications

The C-terminal helix of ribosomal P stalk recognizes a hydrophobic groove of elongation factor 2 in a novel fashion

The C-terminal helix of ribosomal P stalk recognizes a hydrophobic groove of elongation factor 2 in a novel fashion

Eukaryotic elongation factor‐2 (eEF2): its regulation and peptide chain elongation

Control of Ribosomal Subunit Rotation by Elongation Factor G

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