The major long-range goal of this work is obtain high resolution X-ray crystallographic structures of nicotinic acetylcholine receptors (AChRs). This family includes nicotinic acetylcholine (AChR), glycine, gamma- aminobutyric acidA (GABAA), and 5-hydroxytryptamine (5-HT3) receptors. All are involved in signal transmission within the nervous system and have wide-ranging importance in human neurophysiology and neurological diseases. The AChRs are particularly important clinically because of their roles in myasthenia gravis, nicotine addiction, and neurodegenerative diseases. Protein chemistry, ligand binding studies, electrophysiology, and electron microscopy on native and cloned AChRs have combined to elucidate some aspects of the structure and function of these receptors. The crystallographic structures of the AChRs, however, has been elusive. The alpha7 AChR is an attractive candidate for X-ray crystallography of nicotinic receptors because it possesses the simplifying property of containing only the alpha7 type of subunit. The proposed training environment is well-suited to the multidisciplinary nature of this project. The sponsor, Jon Lindstrom is a leader in the biology of AChRs whose laboratory is especially skilled in heterologous expression and functional analysis of AChRs. Patrick loll, an X-ray crystallographer and a collaborator on the project, is experienced with the crystallography of membrane proteins. The methods of protein design, expression and structural analysis complement the previous training of the principle investigator in protein magnetic resonance spectroscopy. They will have general relevance to this entire family ion channels and will be the foundation for his long-range goal of studying the structural biology of ion channels of the nervous system.
