Quinacrine binds to the lipid-protein interface of the Torpedo acetylcholine receptor: a fluorescence study.

Valenzuela, C. Fernando; Kerr, Jennifer; Johnson, David A.

doi:10.1016/s0021-9258(18)42433-7

Cited by 38 publications

(29 citation statements)

References 40 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Inhibition of [I25l]a-BTX Binding to Nicotinic Receptors on Membranes from Torpedo Electric Organ. Membranes containing nAChRs were prepared from Torpedo electric organ and assayed as previously described (Valenzuela et al, 1992;. Torpedo nicotinic receptors (0.3-2.0 nM [125I]a-BTX binding sites) were incubated in 108 pL of assay buffer (10 mM sodium phosphate, pH 7.4, 1.0 mM EDTA, 1.0 mM EGTA, 0.1% Triton X-100) containing metocurine, -conotoxin Ei or Mi, or combinations of the three drugs.…”

Section: Methodsmentioning

confidence: 99%

.alpha.-Conotoxin EI, A New Nicotinic Acetylcholine Receptor Antagonist with Novel Selectivity

Olivera²,

et al. 1995

Biochemistry

103

125

View full text Add to dashboard Cite

We report the isolation and characterization of a novel nicotinic acetylcholine receptor (nAChR) ligand. The toxin is an 18 amino acid peptide and is the first reported alpha-conotoxin from an Atlantic fish-hunting Conus. The peptide was purified from the venom of Conus ermineus and is called alpha-conotoxin EI. The sequence diverges from that of previously isolated alpha-conotoxins. We demonstrate that this structural divergence has functional consequences. In Torpedo nAChRs, alpha-conotoxin EI selectively binds the agonist site near the alpha/delta subunit interface in contrast to alpha-conotoxin MI which selectively targets the alpha/gamma agonist binding site. In mammalian nAChRs alpha-conotoxin EI shows high affinity for both the alpha/delta and alpha/gamma subunit interfaces (with some preference for the alpha/delta site), whereas alpha-conotoxin MI is highly selective for the alpha/delta ligand binding site. The sequence of the peptide is: Arg-Asp-Hyp-Cys-Cys-Tyr-His-Pro-Thr-Cys-Asn-Met-Ser-Asn-Pro-Gln-Ile-Cys- NH2, with disulfide bridging between Cys4-Cys10 and Cys5-Cys18, analogous to those of previously described alpha-conotoxins. This sequence has been verified by total chemical synthesis. Thus, alpha-conotoxin EI is a newly-available tool with unique structure and function for characterization of nAChRs.

show abstract

Section: Methodsmentioning

confidence: 99%

.alpha.-Conotoxin EI, A New Nicotinic Acetylcholine Receptor Antagonist with Novel Selectivity

Olivera²,

et al. 1995

Biochemistry

103

125

View full text Add to dashboard Cite

show abstract

“…where the subscripts D and DA denote the absence and presence of acceptor, respectively. The surface density of acceptor probes was determined in parallel samples by measuring FRET between membrane-partitioned Ci2-fluo-resceinandC|2-eosin or C u-rhodamine as described elsewhere (Valenzuela et al, 1992).…”

Section: Methodsmentioning

confidence: 99%

“…where /pius to* and /minus ,0, ate the magnitudes of fluorescence of samples that did or did not contain an excess of cobra a-toxin, respectively (Valenzuela et al, 1992(Valenzuela et al, , 1994. fa is the magnitude of the intrinsic fluorescence of the samples in the absence of dansyl-C6-choline, and Cd is a correction factor for dilution and inner filter effects of the titrant.…”

Section: Methodsmentioning

confidence: 99%

“…fa is the magnitude of the intrinsic fluorescence of the samples in the absence of dansyl-C6-choline, and Cd is a correction factor for dilution and inner filter effects of the titrant. The correction for inner filter effects was calculated as described by Valenzuela et al (1992). The fraction of nAChR-bound dansyl-CVcholine was determined from a plot of the cobra a-toxin-sensitivc dansyl-C6-choline fluorescence vs the total titrated concentration of dansyl-Cg-choline.…”

Section: Methodsmentioning

confidence: 99%

See 1 more Smart Citation

Antibody-Induced Conformational Changes in the Torpedo Nicotinic Acetylcholine Receptor: A Fluorescence Study

Cf¹,

Aj²,

Hr³

et al. 1994

Biochemistry

View full text Add to dashboard Cite

Quantitative fluorescence spectroscopy was used to develop a structural picture of the effects of two monoclonal antibodies (mAbs) on the conformation of the Torpedo nicotinic acetylcholine receptor (nAChR). The two mAbs (A6 and B1) examined selectively blocked ligand binding to either the high-affinity (A) or the low-affinity (B) binding sites for agonists/competitive antagonists. The distances between dansyl-C6-choline bound to the unblocked agonist/competitive antagonist binding site and one of two lipophilic probes (C12-eosin or C18-rhodamine) partitioned into the lipid membrane were estimated by using fluorescence resonance energy transfer. Control experiments demonstrated that both mAbs decreased the affinity and fluorescence lifetime of receptor-bound dansyl-C6-choline. The binding of the B1 mAb to the B site did not significantly change the calculated distance between the unblocked A binding site and the membrane surface. However, the binding of the A6 mAb to the A site induced the B site to move into close proximity to the lipid membrane. This conformational change was confirmed by a 45-fold increase in the paramagnetic quenching of the B-site-bound dansyl-C6-choline fluorescence by lipid-intercalated 5-doxylstearate. The results indicate that these mAbs not only selectively block ligand binding to either the A or the B acetylcholine sites but also, in the case of the A6 mAb, induce global conformational changes of the receptor, which appear to involve a movement of the B binding site into close proximity of the lipid membrane.(ABSTRACT TRUNCATED AT 250 WORDS)

show abstract

“… 17 In addition, the binding between lipid and quinacrine has reportedly been found to block the Torpedo nicotinic acetylcholine receptor at the lipid–protein interface. 18 , 19 …”

Section: Introductionmentioning

confidence: 99%

Multimodal Investigation into the Interaction of Quinacrine with Microcavity-Supported Lipid Bilayers

2022

View full text Add to dashboard Cite

Quinacrine is a versatile drug that is widely recognized for its antimalarial action through its inhibition of the phospholipase enzyme. It also has antianthelmintic and antiprotozoan activities and is a strong DNA binder that may be used to combat multidrug resistance in cancer. Despite extensive cell-based studies, a detailed understanding of quinacrine’s influence on the cell membrane, including permeability, binding, and rearrangement at the molecular level, is lacking. Herein, we apply microcavity-suspended lipid bilayers (MSLBs) as in vitro models of the cell membrane comprising DOPC, DOPC:Chol(3:1), and DOPC:SM:Chol(2:2:1) to investigate the influence of cholesterol and intrinsic phase heterogeneity induced by mixed-lipid composition on the membrane interactions of quinacrine. Using electrochemical impedance spectroscopy (EIS) and surface-enhanced Raman spectroscopy (SERS) as label-free surface-sensitive techniques, we have studied quinacrine interaction and permeability across the different MSLBs. Our EIS data reveal that the drug is permeable through ternary DOPC:SM:Chol and DOPC-only bilayer compositions. In contrast, the binary cholesterol/DOPC membrane arrested permeation, yet the drug binds or intercalates at this membrane as reflected by an increase in membrane impedance. SERS supported the EIS data, which was utilized to gain structural insights into the drug–membrane interaction. Our SERS data also provides a simple but powerful label-free assessment of drug permeation because a significant SERS enhancement of the drug’s Raman signature was observed only if the drug accessed the plasmonic interior of the pore cavity passing through the membrane. Fluorescent lifetime correlation spectroscopy (FLCS) provides further biophysical insight, revealing that quinacrine binding increases the lipid diffusivity of DOPC and the ternary membrane while remarkably decreasing the lipid diffusivity of the DOPC:Chol membrane. Overall, because of its adaptability to multimodal approaches, the MSLB platform provides rich and detailed insights into drug–membrane interactions, making it a powerful tool for in vitro drug screening.

show abstract

Quinacrine binds to the lipid-protein interface of the Torpedo acetylcholine receptor: a fluorescence study.

Cited by 38 publications

References 40 publications

.alpha.-Conotoxin EI, A New Nicotinic Acetylcholine Receptor Antagonist with Novel Selectivity

.alpha.-Conotoxin EI, A New Nicotinic Acetylcholine Receptor Antagonist with Novel Selectivity

Antibody-Induced Conformational Changes in the Torpedo Nicotinic Acetylcholine Receptor: A Fluorescence Study

Multimodal Investigation into the Interaction of Quinacrine with Microcavity-Supported Lipid Bilayers

Contact Info

Product

Resources

About