scholarly journals Cryo-EM visualization of the ribosome in termination complex with apo-RF3 and RF1

eLife ◽  
2013 ◽  
Vol 2 ◽  
Author(s):  
Jesper Pallesen ◽  
Yaser Hashem ◽  
Gürkan Korkmaz ◽  
Ravi Kiran Koripella ◽  
Chenhui Huang ◽  
...  
Keyword(s):  
G Protein ◽  
Messenger Rna ◽  
Stop Codon ◽  
Transfer Rna ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Rna Translation ◽  
Guanine Nucleotide Exchange ◽  
Messenger Rna Translation ◽  
A Site

Termination of messenger RNA translation in Bacteria and Archaea is initiated by release factors (RFs) 1 or 2 recognizing a stop codon in the ribosomal A site and releasing the peptide from the P-site transfer RNA. After release, RF-dissociation is facilitated by the G-protein RF3. Structures of ribosomal complexes with RF1 or RF2 alone or with RF3 alone—RF3 bound to a non-hydrolyzable GTP-analog—have been reported. Here, we present the cryo-EM structure of a post-termination ribosome containing both apo-RF3 and RF1. The conformation of RF3 is distinct from those of free RF3•GDP and ribosome-bound RF3•GDP(C/N)P. Furthermore, the conformation of RF1 differs from those observed in RF3-lacking ribosomal complexes. Our study provides structural keys to the mechanism of guanine nucleotide exchange on RF3 and to an L12-mediated ribosomal recruitment of RF3. In conjunction with previous observations, our data provide the foundation to structurally characterize the complete action cycle of the G-protein RF3.

scholarly journals Activation of G proteins by guanine nucleotide exchange factors relies on GTPase activity

2015 ◽  
Author(s):  
Rob J Stanley ◽  
Geraint MH Thomas
Keyword(s):  
G Protein ◽  
G Proteins ◽  
Guanine Nucleotide Exchange Factors ◽  
Guanine Nucleotide ◽  
Gtpase Activity ◽  
Nucleotide Exchange ◽  
Signalling Molecules ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factors ◽  
Experimental Strategies

G proteins are an important family of signalling molecules controlled by guanine nucleotide exchange and GTPase activity in what is commonly called an 'activation/inactivation cycle'. The molecular mechanism by which guanine nucleotide exchange factors (GEFs) catalyse the activation of monomeric G proteins is well-established, however the complete reversibility of this mechanism is often overlooked. Here, we use a theoretical approach to prove that GEFs are unable to positively control G protein systems at steady-state in the absence of GTPase activity. Instead, positive regulation of G proteins must be seen as a product of the competition between guanine nucleotide exchange and GTPase activity -- emphasising a central role for GTPase activity beyond merely signal termination. We conclude that a more accurate description of the regulation of G proteins via these processes is as a 'balance/imbalance' mechanism. This result has implications for the understanding of many intracellular signalling processes, and for experimental strategies that rely on modulating G protein systems.

scholarly journals Regulation of the G-protein Regulatory-GαiSignaling Complex by Nonreceptor Guanine Nucleotide Exchange Factors

2012 ◽  
Vol 288 (5) ◽  
pp. 3003-3015 ◽  
Author(s):  
Sukru Sadik Oner ◽  
Ellen M. Maher ◽  
Meital Gabay ◽  
Gregory G. Tall ◽  
Joe B. Blumer ◽  
...  
Keyword(s):  
G Protein ◽  
Guanine Nucleotide Exchange Factors ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factors

Effect of transfer RNA from various sources on placental messenger RNA translation

1983 ◽  
Vol 29 (2) ◽  
pp. 181-195 ◽  
Author(s):  
Susan J. Kelly ◽  
Jean Loria M.T. Gyves ◽  
Judith Ilan
Keyword(s):  
Messenger Rna ◽  
Transfer Rna ◽  
Rna Translation ◽  
Messenger Rna Translation

scholarly journals Drosophila GoLoco-Protein Pins Is a Target of Gαo-mediated G Protein–coupled Receptor Signaling

2009 ◽  
Vol 20 (17) ◽  
pp. 3865-3877 ◽  
Author(s):  
Damir Kopein ◽  
Vladimir L. Katanaev
Keyword(s):  
G Protein ◽  
System Development ◽  
Nervous System Development ◽  
G Protein Coupled Receptors ◽  
Guanine Nucleotide ◽  
Receptor Signaling ◽  
Nucleotide Exchange ◽  
Guanine Nucleotide Exchange ◽  
Necessary And Sufficient ◽  
G Protein Coupled

G protein–coupled receptors (GPCRs) transduce their signals through trimeric G proteins, inducing guanine nucleotide exchange on their Gα-subunits; the resulting Gα-GTP transmits the signal further inside the cell. GoLoco domains present in many proteins play important roles in multiple trimeric G protein–dependent activities, physically binding Gα-subunits of the Gαi/o class. In most cases GoLoco binds exclusively to the GDP-loaded form of the Gα-subunits. Here we demonstrate that the poly-GoLoco–containing protein Pins of Drosophila can bind to both GDP- and GTP-forms of Drosophila Gαo. We identify Pins GoLoco domain 1 as necessary and sufficient for this unusual interaction with Gαo-GTP. We further pinpoint a lysine residue located centrally in this domain as necessary for the interaction. Our studies thus identify Drosophila Pins as a target of Gαo-mediated GPCR receptor signaling, e.g., in the context of the nervous system development, where Gαo acts downstream from Frizzled and redundantly with Gαi to control the asymmetry of cell divisions.

scholarly journals The guanine nucleotide exchange factor Ric-8A induces domain separation and Ras domain plasticity in Gαi1

2015 ◽  
Vol 112 (5) ◽  
pp. 1404-1409 ◽  
Author(s):  
Ned Van Eps ◽  
Celestine J. Thomas ◽  
Wayne L. Hubbell ◽  
Stephen R. Sprang
Keyword(s):  
G Protein ◽  
Bound States ◽  
Guanine Nucleotide Exchange Factor ◽  
Nucleotide Binding ◽  
Conformational Ensemble ◽  
Guanine Nucleotide ◽  
Nucleotide Exchange ◽  
Exchange Factor ◽  
Guanine Nucleotide Exchange ◽  
Nucleotide Exchange Factor

Heterotrimeric G proteins are activated by exchange of GDP for GTP at the G protein alpha subunit (Gα), most notably by G protein-coupled transmembrane receptors. Ric-8A is a soluble cytoplasmic protein essential for embryonic development that acts as both a guanine nucleotide exchange factor (GEF) and a chaperone for Gα subunits of the i, q, and 12/13 classes. Previous studies demonstrated that Ric-8A stabilizes a dynamically disordered state of nucleotide-free Gα as the catalytic intermediate for nucleotide exchange, but no information was obtained on the structures involved or the magnitude of the structural fluctuations. In the present study, site-directed spin labeling (SDSL) together with double electron-electron resonance (DEER) spectroscopy is used to provide global distance constraints that identify discrete members of a conformational ensemble in the Gαi1:Ric-8A complex and the magnitude of structural differences between them. In the complex, the helical and Ras-like nucleotide-binding domains of Gαi1 pivot apart to occupy multiple resolved states with displacements as large as 25 Å. The domain displacement appears to be distinct from that observed in Gαs upon binding of Gs to the β2 adrenergic receptor. Moreover, the Ras-like domain exhibits structural plasticity within and around the nucleotide-binding cavity, and the switch I and switch II regions, which are known to adopt different conformations in the GDP- and GTP-bound states of Gα, undergo structural rearrangements. Collectively, the data show that Ric-8A induces a conformationally heterogeneous state of Gαi and provide insight into the mechanism of action of a nonreceptor Gα GEF.

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