Refolding and characterization of a soluble ectodomain complex of the calcitonin gene-related peptide receptor

Biochemistry
Christopher M KothJonathan M Moore

Abstract

The calcitonin gene-related peptide (CGRP) receptor is a heterodimer of two membrane proteins: calcitonin receptor-like receptor (CLR) and receptor activity-modifying protein 1 (RAMP1). CLR is a class B G-protein-coupled receptor (GPCR), possessing a characteristic large amino-terminal extracellular domain (ECD) important for ligand recognition and binding. Dimerization of CLR with RAMP1 provides specificity for CGRP versus related agonists. Here we report the expression, purification, and refolding of a soluble form of the CGRP receptor comprising a heterodimer of the CLR and RAMP1 ECDs. The extracellular protein domains corresponding to residues 23-133 of CLR and residues 26-117 of RAMP1 were shown to be sufficient for formation of a stable, monodisperse complex. The binding affinity of the purified ECD complex for the CGRP peptide was significantly lower than that of the native receptor (IC(50) of 12 microM for the purified ECD complex vs 233 pM for membrane-bound CGRP receptor), indicating that other regions of CLR and/or RAMP1 are important for peptide agonist binding. However, high-affinity binding to known potent and specific nonpeptide antagonists of the CGRP receptor, including olcegepant and telcagepant (K(D) < 0.02 m...Continue Reading

References

Jan 3, 1985·Nature·S D BrainI MacIntyre
Jan 5, 1995·Biochemical and Biophysical Research Communications·B FlühmannW Born
Nov 1, 1995·Journal of Biomolecular NMR·F DelaglioA Bax
Nov 13, 1998·Proceedings of the National Academy of Sciences of the United States of America·M SalzmannK Wüthrich
Oct 21, 1999·Journal of Magnetic Resonance·J P LoriaA G Palmer
Mar 11, 2000·British Journal of Pharmacology·H DoodsW Eberlein
Feb 16, 2002·The Journal of Biological Chemistry·John J MalleeStefanie A Kane
May 8, 2002·Cephalalgia : an International Journal of Headache·L H LassenJ Olesen
Jun 18, 2002·European Journal of Pharmacology·Marcus Schindler, Henri N Doods
Feb 8, 2003·The Journal of Biological Chemistry·Timothy J FitzsimmonsStephen A Wank
Mar 12, 2004·The New England Journal of Medicine·Jes OlesenUNKNOWN BIBN 4096 BS Clinical Proof of Concept Study Group
Jan 12, 2005·Biochemistry·Madhu ChauhanChandra Yallampalli
Mar 16, 2007·Proceedings of the National Academy of Sciences of the United States of America·Christy Rani R GraceRoland Riek
May 17, 2007·Biochemistry·Madeleine HérouxMichel Bouvier
Jul 17, 2007·Trends in Pharmacological Sciences·Brian K Kobilka, Xavier Deupi
Sep 1, 2007·Nature Methods·Remco SprangersLewis E Kay
Oct 30, 2007·Trends in Pharmacological Sciences·Henri DoodsStefan Just
Jan 25, 2008·The Journal of Headache and Pain·Andrea Stephanie LinkLars Edvinsson
Aug 30, 2008·Protein Science : a Publication of the Protein Society·Seisuke KusanoShigeyuki Yokoyama
Dec 17, 2008·European Journal of Pharmacology·Eric L MooreChristopher A Salvatore

❮ Previous
Next ❯

Citations

Sep 16, 2014·Expert Opinion on Investigational Drugs·László VécseiJános Tajti
May 21, 2010·Expert Opinion on Investigational Drugs·Michael J M Fischer
Oct 31, 2015·Annual Review of Pharmacology and Toxicology·Debbie L Hay, Augen A Pioszak
May 6, 2011·British Journal of Pharmacology·Laurence J MillerKaleeckal G Harikumar
Jan 29, 2013·British Journal of Pharmacology·H A WatkinsD L Hay
Aug 30, 2011·British Journal of Pharmacology·Eric L Moore, Christopher A Salvatore
Oct 23, 2010·Neurogastroenterology and Motility : the Official Journal of the European Gastrointestinal Motility Society·J W RychterA B A Kroese
Oct 12, 2013·Protein Science : a Publication of the Protein Society·Heather E Moad, Augen A Pioszak
Jan 30, 2013·Biochemical Society Transactions·James BarwellDavid R Poyner
Mar 2, 2010·Biochemical and Biophysical Research Communications·E L MooreC A Salvatore
Apr 19, 2017·CNS Neuroscience & Therapeutics·Yogendra SinghKamal Dua
Oct 8, 2014·Physiological Reviews·F A RussellS D Brain
Feb 1, 2012·FASEB Journal : Official Publication of the Federation of American Societies for Experimental Biology·Emilie CeraudoAlain Couvineau
Apr 14, 2016·Biochemical Society Transactions·Debbie L HayDavid R Poyner
Feb 15, 2020·Journal of Medicinal Chemistry·Gene M DubowchikAlison W Xin

❮ Previous
Next ❯

Related Concepts

Trending Feeds

COVID-19

Coronaviruses encompass a large family of viruses that cause the common cold as well as more serious diseases, such as the ongoing outbreak of coronavirus disease 2019 (COVID-19; formally known as 2019-nCoV). Coronaviruses can spread from animals to humans; symptoms include fever, cough, shortness of breath, and breathing difficulties; in more severe cases, infection can lead to death. This feed covers recent research on COVID-19.

Blastomycosis

Blastomycosis fungal infections spread through inhaling Blastomyces dermatitidis spores. Discover the latest research on blastomycosis fungal infections here.

Nuclear Pore Complex in ALS/FTD

Alterations in nucleocytoplasmic transport, controlled by the nuclear pore complex, may be involved in the pathomechanism underlying multiple neurodegenerative diseases including Amyotrophic Lateral Sclerosis and Frontotemporal Dementia. Here is the latest research on the nuclear pore complex in ALS and FTD.

Applications of Molecular Barcoding

The concept of molecular barcoding is that each original DNA or RNA molecule is attached to a unique sequence barcode. Sequence reads having different barcodes represent different original molecules, while sequence reads having the same barcode are results of PCR duplication from one original molecule. Discover the latest research on molecular barcoding here.

Chronic Fatigue Syndrome

Chronic fatigue syndrome is a disease characterized by unexplained disabling fatigue; the pathology of which is incompletely understood. Discover the latest research on chronic fatigue syndrome here.

Evolution of Pluripotency

Pluripotency refers to the ability of a cell to develop into three primary germ cell layers of the embryo. This feed focuses on the mechanisms that underlie the evolution of pluripotency. Here is the latest research.

Position Effect Variegation

Position Effect Variagation occurs when a gene is inactivated due to its positioning near heterochromatic regions within a chromosome. Discover the latest research on Position Effect Variagation here.

STING Receptor Agonists

Stimulator of IFN genes (STING) are a group of transmembrane proteins that are involved in the induction of type I interferon that is important in the innate immune response. The stimulation of STING has been an active area of research in the treatment of cancer and infectious diseases. Here is the latest research on STING receptor agonists.

Microbicide

Microbicides are products that can be applied to vaginal or rectal mucosal surfaces with the goal of preventing, or at least significantly reducing, the transmission of sexually transmitted infections. Here is the latest research on microbicides.