The long-term objectives of this application are to understand how hormones transduce signals to cells. The general strategy is to focus on a superfamily of seven transmembrane-spanning receptors that bind hormones and mediate signals via G proteins. The widespread use of G protein-coupled receptors (GPCRs) as a mechanism of signal transduction into cells make them frequent targets of pharmaceutical drugs; according to one estimate, almost 60% of all drugs act on GPCRs. These include commonly prescribed agents such as beta-blockers (cardiovascular), beta-agonists (asthma), antihistamines (H1- allergies, H2-ulcers), and opiates (pain). An estimated 3% of human genes encode GPCRs; the ligands remain to be identified for nearly half of the receptors, thus offering many new potential drug targets. Despite their biological and medical importance, the molecular mechanisms by which receptors activate G proteins are poorly understood. To address these fundamental questions, this Proposal employs a variety of techniques including genetic screens, computational modeling, and biochemical analyses to study the human complement factor 5 (C5a) receptor, a member of the rhodopsin family of GPCRs. The C5a receptor mediates neutrophil chemotaxis and functions well when expressed in yeast, making possible high-throughput structure/function studies of regions of the receptor that bind hormones or couple to G proteins. Rhodopsin, a photoreceptor activated by light, has been well characterized by biophysical analyses, and is the only GPCR for which a crystal structure is available. The two receptors share similar structures, enabling computational models of the C5a receptor to be generated and for specific models of receptor activation, derived from genetic studies of the C5a receptor, to be tested directly. The studies outlined in this Proposal will provide insights into how receptors function as """"""""on/off"""""""" switches in cells, thus aiding rational drug design for the development of new therapeutic agents. ? ? ?

Agency
National Institute of Health (NIH)
Institute
National Institute of General Medical Sciences (NIGMS)
Type
Research Project (R01)
Project #
1R01GM071634-01A3
Application #
7212648
Study Section
Cellular Signaling and Dynamics Study Section (CSD)
Program Officer
Anderson, Richard A
Project Start
2007-03-01
Project End
2011-02-28
Budget Start
2007-03-01
Budget End
2008-02-29
Support Year
1
Fiscal Year
2007
Total Cost
$260,205
Indirect Cost
Name
Washington University
Department
Other Basic Sciences
Type
Schools of Medicine
DUNS #
068552207
City
Saint Louis
State
MO
Country
United States
Zip Code
63130
Nikiforovich, Gregory V; Baranski, Thomas J (2012) Structural mechanisms of constitutive activation in the C5a receptors with mutations in the extracellular loops: molecular modeling study. Proteins 80:71-80
Nikiforovich, Gregory V; Marshall, Garland R; Baranski, Thomas J (2011) Simplified modeling approach suggests structural mechanisms for constitutive activation of the C5a receptor. Proteins 79:787-802
Nikiforovich, Gregory V; Baranski, Thomas J (2010) Computational modeling of constitutively active mutants of GPCRs C5a receptor. Methods Enzymol 485:369-91
Nikiforovich, Gregory V; Taylor, Christina M; Marshall, Garland R et al. (2010) Modeling the possible conformations of the extracellular loops in G-protein-coupled receptors. Proteins 78:271-85
Van Eps, Ned; Anderson, Lori L; Kisselev, Oleg G et al. (2010) Electron paramagnetic resonance studies of functionally active, nitroxide spin-labeled peptide analogues of the C-terminus of a G-protein alpha subunit. Biochemistry 49:6877-86
Rana, Soumendra; Baranski, Thomas J (2010) Third extracellular loop (EC3)-N terminus interaction is important for seven-transmembrane domain receptor function: implications for an activation microswitch region. J Biol Chem 285:31472-83
Nikiforovich, Gregory V; Baranski, Thomas J (2009) Structural models for the complex of chemotaxis inhibitory protein of Staphylococcus aureus with the C5a receptor. Biochem Biophys Res Commun 390:481-4
Nikiforovich, Gregory V; Marshall, Garland R; Baranski, Thomas J (2008) Modeling molecular mechanisms of binding of the anaphylatoxin C5a to the C5a receptor. Biochemistry 47:3117-30
Hagemann, Ian S; Miller, Daniel L; Klco, Jeffery M et al. (2008) Structure of the complement factor 5a receptor-ligand complex studied by disulfide trapping and molecular modeling. J Biol Chem 283:7763-75