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Title: Targeting G protein-coupled receptor signaling at the G protein level with a selective nanobody inhibitor

Abstract

G protein-coupled receptors (GPCRs) activate heterotrimeric G proteins by mediating a GDP to GTP exchange in the Gα subunit. This leads to dissociation of the heterotrimer into Gα-GTP and Gβγ dimer. The Gα-GTP and Gβγ dimer each regulate a variety of downstream pathways to control various aspects of human physiology. Dysregulated Gβγ-signaling is a central element of various neurological and cancer-related anomalies. However, Gβγ also serves as a negative regulator of Gα that is essential for G protein inactivation, and thus has the potential for numerous side effects when targeted therapeutically. Here we report a llama-derived nanobody (Nb5) that binds tightly to the Gβγ dimer. Nb5 responds to all combinations of β-subtypes and γ-subtypes and competes with other Gβγ-regulatory proteins for a common binding site on the Gβγ dimer. Despite its inhibitory effect on Gβγ-mediated signaling, Nb5 has no effect on Gαq-mediated and Gα s-mediated signaling events in living cells.

Authors:
ORCiD logo [1];  [1];  [2];  [1];  [3]; ORCiD logo [4];  [2];  [5];  [1];  [6]; ORCiD logo [4];  [1]
  1. Case Western Reserve Univ., Cleveland, OH (United States)
  2. The Scripps Research Inst., Jupiter, FL (United States)
  3. Case Western Reserve Univ., Cleveland, OH (United States); Univ. of Colorado School of Medicine, Aurora, CO (United States)
  4. Vrije Universiteit Brussel (VUB) (Belgium); VIB-VUB Center for Structural Biology, Brussels (Belgium)
  5. Case Western Reserve Univ., Cleveland, OH (United States); Louis Stokes Cleveland VA Medical Center, Cleveland, OH (United States)
  6. Univ. of Colorado School of Medicine, Aurora, CO (United States)
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Basic Energy Sciences (BES) (SC-22). Scientific User Facilities Division; National Inst. of Health; Dept. of Veterans Affairs; NIH-ORIP HEI
OSTI Identifier:
1440610
Grant/Contract Number:  
AC02-06CH11357; EY009339; EY027283; EY024864; DA35821; NS95809; P30EY011373; P30EY025585; IK2BX002683; P41 GM103403; S10OD021527
Resource Type:
Journal Article: Accepted Manuscript
Journal Name:
Nature Communications
Additional Journal Information:
Journal Volume: 9; Journal Issue: 1; Journal ID: ISSN 2041-1723
Publisher:
Nature Publishing Group
Country of Publication:
United States
Language:
ENGLISH
Subject:
59 BASIC BIOLOGICAL SCIENCES; Biochemistry; G protein-coupled receptors; Membrane proteins; Structural biology

Citation Formats

Gulati, Sahil, Jin, Hui, Masuho, Ikuo, Orban, Tivadar, Cai, Yuan, Pardon, Els, Martemyanov, Kirill A., Kiser, Philip D., Stewart, Phoebe L., Ford, Christopher P., Steyaert, Jan, and Palczewski, Krzysztof. Targeting G protein-coupled receptor signaling at the G protein level with a selective nanobody inhibitor. United States: N. p., 2018. Web. doi:10.1038/s41467-018-04432-0.
Gulati, Sahil, Jin, Hui, Masuho, Ikuo, Orban, Tivadar, Cai, Yuan, Pardon, Els, Martemyanov, Kirill A., Kiser, Philip D., Stewart, Phoebe L., Ford, Christopher P., Steyaert, Jan, & Palczewski, Krzysztof. Targeting G protein-coupled receptor signaling at the G protein level with a selective nanobody inhibitor. United States. doi:10.1038/s41467-018-04432-0.
Gulati, Sahil, Jin, Hui, Masuho, Ikuo, Orban, Tivadar, Cai, Yuan, Pardon, Els, Martemyanov, Kirill A., Kiser, Philip D., Stewart, Phoebe L., Ford, Christopher P., Steyaert, Jan, and Palczewski, Krzysztof. Fri . "Targeting G protein-coupled receptor signaling at the G protein level with a selective nanobody inhibitor". United States. doi:10.1038/s41467-018-04432-0. https://www.osti.gov/servlets/purl/1440610.
@article{osti_1440610,
title = {Targeting G protein-coupled receptor signaling at the G protein level with a selective nanobody inhibitor},
author = {Gulati, Sahil and Jin, Hui and Masuho, Ikuo and Orban, Tivadar and Cai, Yuan and Pardon, Els and Martemyanov, Kirill A. and Kiser, Philip D. and Stewart, Phoebe L. and Ford, Christopher P. and Steyaert, Jan and Palczewski, Krzysztof},
abstractNote = {G protein-coupled receptors (GPCRs) activate heterotrimeric G proteins by mediating a GDP to GTP exchange in the Gα subunit. This leads to dissociation of the heterotrimer into Gα-GTP and Gβγ dimer. The Gα-GTP and Gβγ dimer each regulate a variety of downstream pathways to control various aspects of human physiology. Dysregulated Gβγ-signaling is a central element of various neurological and cancer-related anomalies. However, Gβγ also serves as a negative regulator of Gα that is essential for G protein inactivation, and thus has the potential for numerous side effects when targeted therapeutically. Here we report a llama-derived nanobody (Nb5) that binds tightly to the Gβγ dimer. Nb5 responds to all combinations of β-subtypes and γ-subtypes and competes with other Gβγ-regulatory proteins for a common binding site on the Gβγ dimer. Despite its inhibitory effect on Gβγ-mediated signaling, Nb5 has no effect on Gαq-mediated and Gαs-mediated signaling events in living cells.},
doi = {10.1038/s41467-018-04432-0},
journal = {Nature Communications},
issn = {2041-1723},
number = 1,
volume = 9,
place = {United States},
year = {2018},
month = {5}
}

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Cited by: 12 works
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    Optical approaches for single-cell and subcellular analysis of GPCR–G protein signaling
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