scholarly journals Activation of GTP hydrolysis in mRNA-tRNA translocation by elongation factor G

2015 ◽  
Vol 1 (4) ◽  
pp. e1500169 ◽  
Author(s):  
Wen Li ◽  
Zheng Liu ◽  
Ravi Kiran Koripella ◽  
Robert Langlois ◽  
Suparna Sanyal ◽  
...  
Keyword(s):  
Elongation Factor ◽  
Elongation Factor G ◽  
Direct Role ◽  
Gtp Hydrolysis ◽  
Ribosomal Subunits ◽  
Trna Translocation ◽  
Gtpase Activation ◽  
Guanosine Triphosphatase ◽  
Factor G

During protein synthesis, elongation of the polypeptide chain by each amino acid is followed by a translocation step in which mRNA and transfer RNA (tRNA) are advanced by one codon. This crucial step is catalyzed by elongation factor G (EF-G), a guanosine triphosphatase (GTPase), and accompanied by a rotation between the two ribosomal subunits. A mutant of EF-G, H91A, renders the factor impaired in guanosine triphosphate (GTP) hydrolysis and thereby stabilizes it on the ribosome. We use cryogenic electron microscopy (cryo-EM) at near-atomic resolution to investigate two complexes formed by EF-G H91A in its GTP state with the ribosome, distinguished by the presence or absence of the intersubunit rotation. Comparison of these two structures argues in favor of a direct role of the conserved histidine in the switch II loop of EF-G in GTPase activation, and explains why GTP hydrolysis cannot proceed with EF-G bound to the unrotated form of the ribosome.

scholarly journals Elongation factor G initiates translocation through a power stroke

2016 ◽  
Vol 113 (27) ◽  
pp. 7515-7520 ◽  
Author(s):  
Chunlai Chen ◽  
Xiaonan Cui ◽  
John F. Beausang ◽  
Haibo Zhang ◽  
Ian Farrell ◽  
...  
Keyword(s):  
Single Molecule ◽  
Messenger Rna ◽  
Elongation Factor ◽  
Power Stroke ◽  
Elongation Factor G ◽  
Gtp Hydrolysis ◽  
Guanosine Triphosphatase ◽  
Prokaryotic Protein ◽  
Domain Iii ◽  
Factor G

During the translocation step of prokaryotic protein synthesis, elongation factor G (EF-G), a guanosine triphosphatase (GTPase), binds to the ribosomal PRE-translocation (PRE) complex and facilitates movement of transfer RNAs (tRNAs) and messenger RNA (mRNA) by one codon. Energy liberated by EF-G’s GTPase activity is necessary for EF-G to catalyze rapid and precise translocation. Whether this energy is used mainly to drive movements of the tRNAs and mRNA or to foster EF-G dissociation from the ribosome after translocation has been a long-lasting debate. Free EF-G, not bound to the ribosome, adopts quite different structures in its GTP and GDP forms. Structures of EF-G on the ribosome have been visualized at various intermediate steps along the translocation pathway, using antibiotics and nonhydolyzable GTP analogs to block translocation and to prolong the dwell time of EF-G on the ribosome. However, the structural dynamics of EF-G bound to the ribosome have not yet been described during normal, uninhibited translocation. Here, we report the rotational motions of EF-G domains during normal translocation detected by single-molecule polarized total internal reflection fluorescence (polTIRF) microscopy. Our study shows that EF-G has a small (∼10°) global rotational motion relative to the ribosome after GTP hydrolysis that exerts a force to unlock the ribosome. This is followed by a larger rotation within domain III of EF-G before its dissociation from the ribosome.

scholarly journals Structure and function of FusB: an elongation factor G-binding fusidic acid resistance protein active in ribosomal translocation and recycling

Open Biology ◽  
2012 ◽  
Vol 2 (3) ◽  
pp. 120016 ◽  
Author(s):  
Xiaohu Guo ◽  
Kristin Peisker ◽  
Kristina Bäckbro ◽  
Yang Chen ◽  
Ravi Kiran Koripella ◽  
...  
Keyword(s):  
Fusidic Acid ◽  
Elongation Factor ◽  
Acid Resistance ◽  
Translation System ◽  
Elongation Factor G ◽  
Gtp Hydrolysis ◽  
Terminal Domain ◽  
Trna Translocation ◽  
And Function ◽  
Factor G

Fusidic acid (FA) is a bacteriostatic antibiotic that locks elongation factor G (EF-G) to the ribosome after GTP hydrolysis during elongation and ribosome recycling. The plasmid pUB101-encoded protein FusB causes FA resistance in clinical isolates of Staphylococcus aureus through an interaction with EF-G. Here, we report 1.6 and 2.3 Å crystal structures of FusB. We show that FusB is a two-domain protein lacking homology to known structures, where the N-terminal domain is a four-helix bundle and the C-terminal domain has an alpha/beta fold containing a C4 treble clef zinc finger motif and two loop regions with conserved basic residues. Using hybrid constructs between S. aureus EF-G that binds to FusB and Escherichia coli EF-G that does not, we show that the sequence determinants for FusB recognition reside in domain IV and involve the C-terminal helix of S. aureus EF-G. Further, using kinetic assays in a reconstituted translation system, we demonstrate that FusB can rescue FA inhibition of tRNA translocation as well as ribosome recycling. We propose that FusB rescues S. aureus from FA inhibition by preventing formation or facilitating dissociation of the FA-locked EF-G–ribosome complex.

scholarly journals Activation of GTP hydrolysis in mRNA-tRNA translocation by elongation factor G

2018 ◽  
pp. 490-496
Author(s):  
Wen Li ◽  
Zheng Liu ◽  
Ravi Kiran Koripella ◽  
Robert Langlois ◽  
Suparna Sanyal ◽  
...  
Keyword(s):  
Elongation Factor ◽  
Elongation Factor G ◽  
Gtp Hydrolysis ◽  
Trna Translocation ◽  
Factor G

scholarly journals Active role of elongation factor G in maintaining the mRNA reading frame during translation

2019 ◽  
Vol 5 (12) ◽  
pp. eaax8030 ◽  
Author(s):  
Bee-Zen Peng ◽  
Lars V. Bock ◽  
Riccardo Belardinelli ◽  
Frank Peske ◽  
Helmut Grubmüller ◽  
...  
Keyword(s):  
Elongation Factor ◽  
Active Role ◽  
Transfer Rna ◽  
Specific Interactions ◽  
Elongation Factor G ◽  
Reading Frame ◽  
A Site ◽  
Factor G ◽  
Domain 4

During translation, the ribosome moves along the mRNA one codon at a time with the help of elongation factor G (EF-G). Spontaneous changes in the translational reading frame are extremely rare, yet how the precise triplet-wise step is maintained is not clear. Here, we show that the ribosome is prone to spontaneous frameshifting on mRNA slippery sequences, whereas EF-G restricts frameshifting. EF-G helps to maintain the mRNA reading frame by guiding the A-site transfer RNA during translocation due to specific interactions with the tip of EF-G domain 4. Furthermore, EF-G accelerates ribosome rearrangements that restore the ribosome’s control over the codon-anticodon interaction at the end of the movement. Our data explain how the mRNA reading frame is maintained during translation.

scholarly journals Allosteric collaboration between elongation factor G and the ribosomal L1 stalk directs tRNA movements during translation

2009 ◽  
Vol 106 (37) ◽  
pp. 15702-15707 ◽  
Author(s):  
Jingyi Fei ◽  
Jonathan E. Bronson ◽  
Jake M. Hofman ◽  
Rathi L. Srinivas ◽  
Chris H. Wiggins ◽  
...  
Keyword(s):  
Dynamic Equilibrium ◽  
Ribosomal Subunit ◽  
Elongation Factor ◽  
Large Ribosomal Subunit ◽  
Elongation Factor G ◽  
Structural Element ◽  
Time Dynamics ◽  
Kinetic Mechanisms ◽  
Trna Translocation ◽  
Factor G

Determining the mechanism by which tRNAs rapidly and precisely transit through the ribosomal A, P, and E sites during translation remains a major goal in the study of protein synthesis. Here, we report the real-time dynamics of the L1 stalk, a structural element of the large ribosomal subunit that is implicated in directing tRNA movements during translation. Within pretranslocation ribosomal complexes, the L1 stalk exists in a dynamic equilibrium between open and closed conformations. Binding of elongation factor G (EF-G) shifts this equilibrium toward the closed conformation through one of at least two distinct kinetic mechanisms, where the identity of the P-site tRNA dictates the kinetic route that is taken. Within posttranslocation complexes, L1 stalk dynamics are dependent on the presence and identity of the E-site tRNA. Collectively, our data demonstrate that EF-G and the L1 stalk allosterically collaborate to direct tRNA translocation from the P to the E sites, and suggest a model for the release of E-site tRNA.

Crystallographic studies of elongation factor G

1995 ◽  
Vol 73 (11-12) ◽  
pp. 1209-1216 ◽  
Author(s):  
Anders Liljas ◽  
Arnthor Ævarsson ◽  
Salam Al-Karadaghi ◽  
Maria Garber ◽  
Julia Zheltonosova ◽  
...  
Keyword(s):  
Crystal Structures ◽  
Conformational Change ◽  
Conformational Changes ◽  
Elongation Factor ◽  
Elongation Factors ◽  
Elongation Factor G ◽  
Conformational States ◽  
Gtp Hydrolysis ◽  
Large Conformational Change ◽  
Factor G

The elongation factors G (EF-G) and Tu (EF-Tu) go through a number of conformation states in their functional cycles. Since they both are GTPases, have similar G domains and domains II, and have similar interactions with the nucleotides, then GTP hydrolysis must occur in similar ways. The crystal structures of two conformational states are known for EF-G and three are known for EF-Tu. The conformations of EF-G∙GDP and EF-Tu∙GTP are closely related. EF-Tu goes through a large conformational change upon GTP cleavage. This conformational change is to a large extent due to an altered interaction between the G domain and domains II and III. A number of kirromycin-resistant mutations are situated at the interface between domains I and III. The interface between the G domain and domain V in EF-G corresponds with this dynamic interface in EF-Tu. The contact area in EF-G is small and dominated by interactions between charged amino acids, which are part of a system that is observed to undergo conformational changes. Furthermore, a number of fusidic acid resistant mutants have been identified in this area. All of this evidence makes it likely that EF-G undergoes a large conformational change in its functional cycle. If the structures and conformational states of the elongation factors are related to a scheme in which the ribosome oscillates between two conformations, the pretranslocational and posttranslocational states, a model is arrived at in which EF-Tu drives the reaction in one direction and EF-G in the opposite. This may lead to the consequence that the GTP state of one factor is similar to the GDP state of the other. At the GTP hydrolysis state, the structures of the factors will be close to superimposable.Key words: elongation factor G, elongation factor Tu, crystal structures, conformational changes, ribosomal conformation.

scholarly journals Control of Ribosomal Subunit Rotation by Elongation Factor G

Science ◽  
2013 ◽  
Vol 340 (6140) ◽  
pp. 1235970 ◽  
Author(s):  
Arto Pulk ◽  
Jamie H. D. Cate
Keyword(s):  
Messenger Rna ◽  
Ribosomal Subunit ◽  
Elongation Factor ◽  
Peptide Bond ◽  
Elongation Factor G ◽  
Before And After ◽  
Switch Regions ◽  
Subunit Rotation ◽  
Guanosine Triphosphatase ◽  
Factor G

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 angstrom resolution x-ray crystal structures of EF-G complexed with a nonhydrolyzable guanosine 5′-triphosphate (GTP) analog 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 before and after GTP hydrolysis.

Sign in / Sign up

Export Citation Format

Share Document