Correlation between acetylcholine receptor function and structural properties of membranes

Abstract

Protein-lipid interactions were studied by using Torpedo californica acetylcholine receptor (AChR) as a model system by reconstituting purified AChR into membranes containing various synthetic lipids and native lipids. AChR function was determined by measuring two activities at 4 degrees C: (1) low to high agonist affinity-state transition of AChR in the presence of an agonist (carbamylcholine) in either membrane fragments or sealed vesicles and (2) ion-gating activity of AChR-containing vesicles in response to carbamylcholine. Sixteen samples were examined, each containing different lipid compositions including phosphatidylcholine, cholesterol, phosphatidic acid, phosphatidylethanolamine, asolectin, neutral lipid depleted asolectin, native lipids, and cholesterol-depleted native lipids. Phosphatidylcholines with different configurations of fatty acyl chains were used. The dynamic structures of these membranes were probed by incorporating spin-labeled fatty acid into AChR-containing vesicles and measuring the order parameters. It was found that both aspects of AChR function were highly dependent on the lipid environment even though carbamylcholine binding itself was not affected. An appropriate membrane fluidity was necessarily required to allow the interconversion between the low and high affinity states of AChR. An optimal fluidity hypothesis is proposed to account for the conformational transition properties of membrane proteins. In addition, the conformational change was only a necessary, but not sufficient, condition for the AChR-mediated ion flux activity. Among membranes in which AChR manifested the affinity-state transition, only those containing both cholesterol and negatively charged phospholipids (such as phosphatidic acid) retained the ion-gating activity.

Structural Basis for Allosteric Coupling at the Membrane-Protein Interface in Gloeobacter violaceus Ligand-gated Ion Channel (GLIC)

“…There is also growing evidence that crystallization conditions may bias structures toward specific conformations (6,11,63). Gating in these channels is intrinsically modulated by membrane lipids (22,64,65), likely by an allosteric mechanism that alters the relay of communication through the interface between the ECD and the TMD. Furthermore, it is also likely that these channels from different phylogenetic origins may have intrinsically different mechanisms.…”

[3H]Epibatidine Photolabels Non-equivalent Amino Acids in the Agonist Binding Site of Torpedo and α4β2 Nicotinic Acetylcholine Receptors

“…Torpedo nAChR-rich membranes at 1 mg/ml protein were solubilized in 1% sodium cholate in vesicle dialysis buffer (VDB; 100 mM NaCl, 0.1 mM EDTA, 0.02% NaN 3 , 10 mM MOPS, pH 7.5) and treated with 0.1 mM diisopropylfluorophosphate after insoluble material was pelleted by centrifugation (91,000 ϫ g for 1 h). The nAChR was affinity-purified on a bromoacetylcholine bromide-derivatized Affi-Gel 10 column and then reconstituted into lipid vesicles composed of dioleoyl phosphatidic acid/dioleoyl phosphatidylcholine/cholesterol (at a molar ratio of 3:1:1), as described (23,24). The lipid/nAChR ratio was adjusted to molar ratio of 400:1.…”

Acetyl-L-carnitine physical-chemical, metabolic, and therapeutic properties: relevance for its mode of action in Alzheimer's disease and geriatric depression

“…[143][144][145][146][147][148][149][150][151][152][153] Since many membrane receptors and channels contain several transmembrane helical regions, 154,155 the surrounding membrane phospholipids are poised to influence the structure and molecular dynamics of the transmembrane regions of receptors and ion channels. [156][157][158][159][160][161][162] Membrane molecular dynamics also could influence the coupling of G-protein subunits to receptor subunits, the coupling of G-protein subunits to each other, and the coupling of G-protein subunits to membrane associated enzymes such as adenylcyclase and phospholipase C.…”