title: Agonist-mediated conformational changes in acetylcholine-binding protein revealed by simulation and intrinsic tryptophan fluorescence.
abstract: We delineated acetylcholine (ACh)-dependent conformational changes in a prototype of the nicotinic receptor ligand binding domain by molecular dynamics simulation and changes in intrinsic tryptophan (Trp) fluorescence. Prolonged molecular dynamics simulation of ACh-binding protein showed that binding of ACh establishes close register of Trps from adjacent subunits, Trp(143) and Trp(53), and draws the peripheral C-loop inward to occlude the entrance to the binding cavity. Close register of Trp(143) and Trp(53) was demonstrated by ACh-mediated quenching of intrinsic Trp fluorescence, elimination of quenching by mutation of one or both Trps to Phe, and decreased lifetime of Trp fluorescence by bound ACh. Occlusion of the binding cavity by the C-loop was demonstrated by restricted access of an extrinsic quencher of binding site Trp fluorescence by ACh. The collective findings showed that ACh initially establishes close register of conserved Trps from adjacent subunits and then draws the C-loop inward to occlude the entrance to the binding cavity.
authors.affiliation: Receptor Biology Laboratory, Department of Physiology and Biomedical Engineering, Mayo Clinic College of Medicine, Rochester, Minnesota 55905, USA.
authors.forename: Fan
authors.initials: F
authors.lastname: Gao
authors.forename: Nina
authors.initials: N
authors.lastname: Bren
authors.forename: Thomas P
authors.initials: TP
authors.lastname: Burghardt
authors.forename: Scott
authors.initials: S
authors.lastname: Hansen
authors.forename: Richard H
authors.initials: RH
authors.lastname: Henchman
authors.forename: Palmer
authors.initials: P
authors.lastname: Taylor
authors.forename: J Andrew
authors.initials: JA
authors.lastname: McCammon
authors.forename: Steven M
authors.initials: SM
authors.lastname: Sine
mesh_terms: D000109:Acetylcholine / Q000737:chemistry*; D020106:Acrylamide / Q000737:chemistry; D000818:Animals; D001665:Binding Sites; D002352:Carrier Proteins / Q000819:agonists* / Q000737:chemistry*; D018360:Crystallography, X-Ray; D006801:Humans; D008024:Ligands; D008958:Models, Molecular; D011485:Protein Binding; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D012908:Snails; D013050:Spectrometry, Fluorescence; D013997:Time Factors; D014364:Tryptophan / Q000737:chemistry*
chemical_list: C428944:AChBP protein, Lymnaea; D002352:Carrier Proteins; D008024:Ligands; D020106:Acrylamide; D014364:Tryptophan; D000109:Acetylcholine