Nicotinic acetylcholine receptors and GABAA receptors are members of the nicotinoid family of ligand-gated ion channel receptors of the central nervous system. They play major roles in addiction to nicotine, cocaine, alcohol, and barbiturates. Better prevention or treatment of these addictions will have major impact on human heath. High-resolution structures of these receptors are needed as a foundation on for understanding how these receptors work, how they affect addition, and how receptor-specific drugs can be designed for targeted therapeutics for addiction and also for Alzheimer's and Parkinson's diseases, epilepsy, and mental illnesses. The acetylcholine binding protein is limited as a structural model of nicotinoid receptors, because it is not an ion channel. Structures of functional nicotinoid receptors, however, have been elusive. A major obstacle has been the difficulty of obtaining enough protein, either for X-ray crystallography or with isotopic labeling for NMR. Although no bacterial nicotinoid receptors have yet been identified, such bacterial receptors could, in principle, solve this problem. Similar to the impact that bacterial potassium and chloride ion channels have on understanding those families of channels, bacterial nicotinoid receptors promise to significantly advance the understanding of the structure and function of nicotinoid receptors. We have identified several bacterial proteins whose primary amino acid sequences are similar to the extracellular domain and transmembrane domains of nicotinic acetylcholine, ionotropic GABA, and glycine receptors. The long-range goal of this project is to develop an atomic-level structural interpretation of how nicotinoid receptors contribute to addiction. The objective of this application is to determine whether bacterial proteins belong to the nicotinoid receptor family. The central hypothesis is that bacterial proteins with primary amino acid sequences that are similar to eukaryotic nicotinoid subunits are members of the nicotinoid receptor family.
These specific aims test this hypothesis: (1) determine whether bacterial proteins with substantial primary sequence homology to subunits in nicotinoid family assemble into oligomers; (2) determine whether these proteins are transported to the cell surface; and (3) determine electrophysiological properties of these proteins. Identifying the first bacterial nicotinoid receptors will provide promising candidates for structural study of this important family of receptors.
