- Core C (Protein Chemistry Core) The Protein Chemistry Core will continue to provide the research groups in the Program Project with the technical expertise and equipment required for the isolation and sequence analysis of receptor fragments containing the sites of photoincorporation by general anesthetics. In addition, the Core provides computational resources for the homology modeling and docking studies that are necessary to interpret the protein chemistry results in terms of models of receptor structure (Projects 1 and 2) and to provide guidance in the design and interpretation of mutational analyses (Project 3) that further assess the structure and function of the identified binding sites. The Core is under the supervision of Dr. Cohen and is located in his laboratory. Major equipment currently available in the Core include (i) an Applied Biosystems 492 Procise automated Protein Sequenator and in-line amino acid analyzer;and (ii) 2 Agilent 1100 HPLCs equipped with UV and fluorescence detectors;and (iii) an Agilent 1100 capillary LC system. The biochemical characterization of anesthetic binding sites in ligand-gated ion channels requires the use of highly specialized techniques not normally available in protein chemistry core facilities which usually do not accept radioactive samples either for Edman degradation or mass spectrometry. The identification of drug binding sites within the hydrophobic domains of integral membrane proteins poses unique problems for peptide isolation and characterization by either Edman degradation or mass spectrometry. It is the function of the Core to provide the appropriate equipment and highly skilled protein chemists who can interact directly with the research projects to develop and carry out the appropriate research strategies and to educate the investigators about the necessary protein chemistry techniques to be carried out either in the Core or in the investigator's lab. In the previous funding period, novel experimental strategies were developed to characterize anesthetic photolabeling of amino acids in each of the transmembrane helices of the a1?3g2 GABAAR, which led to the identification of a second class of GABAAR anesthetic binding sites, and in serotonin 5-HT3 receptors. This Core will now provide similar expertise for the characterization of novel anesthetic binding sites in a1?3g2 and a4?3d GABAARs and GlyRs.

National Institute of Health (NIH)
Research Program Projects (P01)
Project #
Application #
Study Section
Special Emphasis Panel (ZGM1)
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
Massachusetts General Hospital
United States
Zip Code
Forman, Stuart A; Chiara, David C; Miller, Keith W (2015) Anesthetics target interfacial transmembrane sites in nicotinic acetylcholine receptors. Neuropharmacology 96:169-77
Feng, H-J; Jounaidi, Y; Haburcak, M et al. (2014) Etomidate produces similar allosteric modulation in *1*3ýý and *1*3ýý2L GABA(A) receptors. Br J Pharmacol 171:789-98
Jayakar, Selwyn S; Zhou, Xiaojuan; Chiara, David C et al. (2014) Multiple propofol-binding sites in a ?-aminobutyric acid type A receptor (GABAAR) identified using a photoreactive propofol analog. J Biol Chem 289:27456-68
Dostalova, Zuzana; Zhou, Xiaojuan; Liu, Aiping et al. (2014) Human ?1?3?2L gamma-aminobutyric acid type A receptors: High-level production and purification in a functional state. Protein Sci 23:157-66
Olsen, Richard W; Li, Guo-Dong; Wallner, Martin et al. (2014) Structural models of ligand-gated ion channels: sites of action for anesthetics and ethanol. Alcohol Clin Exp Res 38:595-603
Hamouda, Ayman K; Stewart, Deirdre S; Chiara, David C et al. (2014) Identifying barbiturate binding sites in a nicotinic acetylcholine receptor with [3H]allyl m-trifluoromethyldiazirine mephobarbital, a photoreactive barbiturate. Mol Pharmacol 85:735-46
Chiara, David C; Gill, Jonathan F; Chen, Qiang et al. (2014) Photoaffinity labeling the propofol binding site in GLIC. Biochemistry 53:135-42
Jayakar, Selwyn S; Dailey, William P; Eckenhoff, Roderic G et al. (2013) Identification of propofol binding sites in a nicotinic acetylcholine receptor with a photoreactive propofol analog. J Biol Chem 288:6178-89
Stewart, Deirdre S; Hotta, Mayo; Desai, Rooma et al. (2013) State-dependent etomidate occupancy of its allosteric agonist sites measured in a cysteine-substituted GABAA receptor. Mol Pharmacol 83:1200-8
Forman, Stuart A (2013) A paradigm shift from biophysical to neurobiological: the fading influence of Claude Bernard's ideas about general anesthesia. Anesthesiology 118:984-5

Showing the most recent 10 out of 85 publications