期刊
INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES
卷 41, 期 4, 页码 423-429出版社
ELSEVIER
DOI: 10.1016/j.ijbiomac.2007.05.012
关键词
human nicotinic acetylcholine receptor; extracellular domain; circular dichroism spectroscopy; ligand-binding; secondary and tertiary structure
The extracellular domains (ECDs) of human nicotinic acetylcholine receptors (nAChRs) are of major pharmacological interest as drug targets in the autoimmune disease myasthenia gravis and in various neurological disorders. We have previously expressed and purified the human muscle alpha 1-, beta 1-, gamma- and epsilon-nAChR-ECDs, as well as the wild type and a mutant of neuronal alpha 7-ECD, in yeast Pichia pastoris. The far-UV circular dichroism (CD) studies of these ECDs, presented here, revealed a major prevalence of beta-sheet (similar to 40%) and a small proportion of alpha-helical (similar to 5%) structure for all ECDs, in good agreement with the secondary structure composition of the Torpedo muscle-type nAChR-ECDs and in less, but considerable, agreement with that of the homologous invertebrate acetylcholine-binding proteins (AChBPs). The near-UV CD studies of these nAChR-ECDs indicated well-defined tertiary structures, as was previously suggested by biochemical and immunochemical studies. Furthermore, the binding of cholinergic ligands to the mutant of alpha 7-ECD resulted in no changes in its secondary structure, but revealed significant local conformational changes. Our present studies probe the structure of human nAChR-ECDs for the first time and further suggest that our expressed proteins fold to a near-native conformation, thus being suitable for further structural studies. (C) 2007 Elsevier B.V. All rights reserved.
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