The recent discoveries that opioid receptors, which are members of the G-protein coupled receptor (GPCR) family, can exist as heteromers in live cells, and that 8-|x and 6-K opioid receptor complexes are distinct functional signaling units, have added a new dimension of complexity to the opioid research field. Structural and mechanistic information about opioid receptor dimeric/oligomeric complexes has therefore become of major importance for understanding the mechanisms of action of opiates. The goal of the work proposed in this grant application is to identify the molecular determinants responsible for the oligomerization of 8- and |>opioid receptors (both homo- and heteromers) in a structural context of receptor models, using an iterative combined computational and experimental approach. Since there is no detailed structural data about opioid receptor dimers, and no evidence that the homo- and hetero-oligomerization interfaces of GPCRs coincide, we developed two different computational approaches based on correlated mutation analysis, and identified the most likely homo- and hetero- dimerization interfaces between the transmembrane regions of opioid receptor subtypes. Inferences from these bioinformatics tools and initial molecular models obtained in the preliminary studies constitute the pilot data for the combined computational and experimental strategy that will serve, as described in this grant application, to probe, validate, and refine models of 8- and |i-opioid receptor homo- and heteromers. An important goal of these studies is to be able to identify specific mutations that can be used to modulate dimerization/oligomerization, and thus affect receptor function in a manner that will reveal physiologically relevant mechanisms that depend and/or ensue from dimerization. The combined computational and experimental strategy proposed in this grant application is also expected to provide direct evidence either for the formation of subtype-specific interface(s) in 8- and |i- opioid receptor dimers, or for the arrangement of these receptors into higher-order oligomers with distinct symmetrical or asymmetrical interfaces, as recently proposed for rhodopsin oligomers. Inferences from these studies are expected to provide new insights into the mechanisms underlying opioid-receptor function, with the ultimate goal of helping drug design. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute on Drug Abuse (NIDA)
Type
Research Project (R01)
Project #
1R01DA020032-01A1
Application #
7103043
Study Section
Molecular Neuropharmacology and Signaling Study Section (MNPS)
Program Officer
Hillery, Paul
Project Start
2006-03-01
Project End
2011-02-28
Budget Start
2006-03-01
Budget End
2007-02-28
Support Year
1
Fiscal Year
2006
Total Cost
$325,722
Indirect Cost
Name
Weill Medical College of Cornell University
Department
Physiology
Type
Schools of Medicine
DUNS #
060217502
City
New York
State
NY
Country
United States
Zip Code
10065
Johnston, Jennifer M; Wang, Hao; Provasi, Davide et al. (2012) Assessing the relative stability of dimer interfaces in g protein-coupled receptors. PLoS Comput Biol 8:e1002649
Johnston, Jennifer M; Filizola, Marta (2011) Showcasing modern molecular dynamics simulations of membrane proteins through G protein-coupled receptors. Curr Opin Struct Biol 21:552-8
Johnston, Jennifer M; Aburi, Mahalaxmi; Provasi, Davide et al. (2011) Making structural sense of dimerization interfaces of delta opioid receptor homodimers. Biochemistry 50:1682-90
Golebiewska, Urszula; Johnston, Jennifer M; Devi, Lakshmi et al. (2011) Differential response to morphine of the oligomeric state of ýý-opioid in the presence of ýý-opioid receptors. Biochemistry 50:2829-37
Provasi, Davide; Filizola, Marta (2010) Putative active states of a prototypic g-protein-coupled receptor from biased molecular dynamics. Biophys J 98:2347-55
Filizola, Marta (2010) Increasingly accurate dynamic molecular models of G-protein coupled receptor oligomers: Panacea or Pandora's box for novel drug discovery? Life Sci 86:590-7
Provasi, Davide; Johnston, Jennifer M; Filizola, Marta (2010) Lessons from free energy simulations of delta-opioid receptor homodimers involving the fourth transmembrane helix. Biochemistry 49:6771-6
Mobarec, Juan Carlos; Sanchez, Roberto; Filizola, Marta (2009) Modern homology modeling of G-protein coupled receptors: which structural template to use? J Med Chem 52:5207-16
Provasi, Davide; Bortolato, Andrea; Filizola, Marta (2009) Exploring molecular mechanisms of ligand recognition by opioid receptors with metadynamics. Biochemistry 48:10020-9
Bortolato, A; Mobarec, J C; Provasi, D et al. (2009) Progress in elucidating the structural and dynamic character of G Protein-Coupled Receptor oligomers for use in drug discovery. Curr Pharm Des 15:4017-25

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