Identification of a fluorescent general anesthetic, 1-aminoanthracene

Butts, Christopher A.; Jin, Xi; Brannigan, Grace; Saad, Abdalla A.; Venkatachalan, Srinivasan P; Pearce, Robert A.; Klein, Michael L.; Eckenhoff, Roderic G.; Dmochowski, Ivan J.

doi:10.1073/pnas.0810590106

Cited by 47 publications

(85 citation statements)

References 36 publications

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“…In addition, apoferritin is commercially available and crystallizes readily. Most importantly, however, the affinity of HSAF for a broad range of general anesthetics is highly correlated with anesthetic potency, confirming the utility and relevance of this model system (17).…”

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confidence: 58%

A Unitary Anesthetic Binding Site at High Resolution

Vedula¹,

Brannigan

Economou

et al. 2009

Journal of Biological Chemistry

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156

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Propofol is the most widely used injectable general anesthetic. Its targets include ligand-gated ion channels such as the GABA A receptor, but such receptor-channel complexes remain challenging to study at atomic resolution. Until structural biology methods advance to the point of being able to deal with systems such as the GABA A receptor, it will be necessary to use more tractable surrogates to probe the molecular details of anesthetic recognition. We have previously shown that recognition of inhalational general anesthetics by the model protein apoferritin closely mirrors recognition by more complex and clinically relevant protein targets; here we show that apoferritin also binds propofol and related GABAergic anesthetics, and that the same binding site mediates recognition of both inhalational and injectable anesthetics. Apoferritin binding affinities for a series of propofol analogs were found to be strongly correlated with the ability to potentiate GABA responses at GABA A receptors, validating this model system for injectable anesthetics. High resolution x-ray crystal structures reveal that, despite the presence of hydrogen bond donors and acceptors, anesthetic recognition is mediated largely by van der Waals forces and the hydrophobic effect. Molecular dynamics simulations indicate that the ligands undergo considerable fluctuations about their equilibrium positions. Finally, apoferritin displays both structural and dynamic responses to anesthetic binding, which may mimic changes elicited by anesthetics in physiologic targets like ion channels.Most general anesthetics alter the activity of ligand-gated ion channels, and electrophysiology, photolabeling, and transgenic animal experiments imply that this effect contributes to the mechanism of anesthesia (1-9). Although the molecular mechanism for this effect is not yet clear, photolabeling studies indicate that anesthetics bind within the transmembrane regions of Cys-loop ligand-gated ion channels such as the nicotinic acetylcholine and the ␥-aminobutyric acid (GABA) 2 type A receptors (2, 9 -11). Practical difficulties associated with overexpression, purification, and crystallization of ion channels have thus far stymied investigation of the structural and energetic bases underlying anesthetic recognition. However, general anesthetics also bind specifically to sites in soluble proteins, including firefly luciferase, human serum albumin (HSA), and horse spleen apoferritin (HSAF) (12)(13)(14), and x-ray crystal structures have been determined for complexes of these proteins with several general anesthetics (14 -16). In particular, HSAF is an attractive model for studying anesthetic-protein interactions because it has the highest affinity for anesthetics of any protein studied to date, has a unique anesthetic binding site, and is a multimer of 4-helix bundles, much like the putative anesthetic binding regions in ligand-gated channels. In addition, apoferritin is commercially available and crystallizes readily. Most importantly, however, the affinity of HSAF f...

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mentioning

confidence: 58%

A Unitary Anesthetic Binding Site at High Resolution

Vedula¹,

Brannigan

Economou

et al. 2009

Journal of Biological Chemistry

Self Cite

156

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“…Secondly, no ordered SDS molecules are seen to be adsorbed onto the apoferritin outer or inner protein surfaces; instead, only a single SDS molecule binds in an internal cavity, the same site that has been shown to bind many general anesthetics, including both inhaled and intravenous agents ( Fig. 5; Butts et al, 2009;Vedula et al, 2009). This site has also been reported to bind other amphipathic molecules (Niemeyer et al, 2008;Trikha et al, 1995).…”

Section: Discussionmentioning

confidence: 95%

Beyond the detergent effect: a binding site for sodium dodecyl sulfate (SDS) in mammalian apoferritin

Liu

Jin

et al. 2012

Acta Crystallogr D Biol Cryst

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Although sodium dodecyl sulfate (SDS) is widely used as an anionic detergent, it can also exert specific pharmacological effects that are independent of the surfactant properties of the molecule. However, structural details of how proteins recognize SDS are scarce. Here, it is demonstrated that SDS binds specifically to a naturally occurring four-helix bundle protein: horse apoferritin. The X-ray crystal structure of the apoferritin-SDS complex was determined at a resolution of 1.9 Å and revealed that the SDS binds in an internal cavity that has previously been shown to recognize various general anesthetics. A dissociation constant of 24 AE 9 mM at 293 K was determined by isothermal titration calorimetry. SDS binds in this cavity by bending its alkyl tail into a horseshoe shape; the charged SDS head group lies in the opening of the cavity at the protein surface. This crystal structure provides insights into the protein-SDS interactions that give rise to binding and may prove useful in the design of novel SDS-like ligands for some proteins.

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“…Thus, we assume that the commonly used medical radioisotopes such as gallium-67, rubidium-82, copper-64, technetium-99m, indium-111, xenon-133, thallium-201, and yttrium-90 can also be loaded into HFn nanocages for tumor-targeted therapy and diagnosis. So far, a great number of foreign molecules have been successfully encapsulated into ferritin nanocages (18)(19)(20)(21)(22)(23)(24), indicating that HFn is a universal drug carrier for tumor-targeted delivery.…”

Section: Easy Scaling-up and Manufacturing With Robust And Reproduciblementioning

confidence: 99%

“…In the present study, we loaded HFn nanocage with doxorubicin (Dox) for tumor-specific drug delivery. HFn nanocages can encapsulate large amounts of foreign molecules (18)(19)(20)(21)(22)(23)(24), bind specifically to tumor cells that overexpress TfR1 (17), and should be able to efficiently deliver high doses of therapeutic drugs to tumors. In particular, natural HFn nanocarriers are expected to possess an outstanding biocompatibility and safety profile, because they exist naturally in the human body and are composed of nontoxic elements that therefore would not activate inflammatory or immunological responses (25).…”

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confidence: 99%

H-ferritin–nanocaged doxorubicin nanoparticles specifically target and kill tumors with a single-dose injection

Liang

Fan

Zhou

et al. 2014

Proc. Natl. Acad. Sci. U.S.A.

412

438

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An ideal nanocarrier for efficient drug delivery must be able to target specific cells and carry high doses of therapeutic drugs and should also exhibit optimized physicochemical properties and biocompatibility. However, it is a tremendous challenge to engineer all of the above characteristics into a single carrier particle. Here, we show that natural H-ferritin (HFn) nanocages can carry high doses of doxorubicin (Dox) for tumor-specific targeting and killing without any targeting ligand functionalization or property modulation. Doxloaded HFn (HFn-Dox) specifically bound and subsequently internalized into tumor cells via interaction with overexpressed transferrin receptor 1 and released Dox in the lysosomes. In vivo in the mouse, HFn-Dox exhibited more than 10-fold higher intratumoral drug concentration than free Dox and significantly inhibited tumor growth after a single-dose injection. Importantly, HFn-Dox displayed an excellent safety profile that significantly reduced healthy organ drug exposure and improved the maximum tolerated dose by fourfold compared with free Dox. Moreover, because the HFn nanocarrier has well-defined morphology and does not need any ligand modification or property modulation it can be easily produced with high purity and yield, which are requirements for drugs used in clinical trials. Thus, these unique properties make the HFn nanocage an ideal vehicle for efficient anticancer drug delivery.

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Identification of a fluorescent general anesthetic, 1-aminoanthracene

Abstract: We identified a fluorophore, 1-aminoanthracene (1-AMA), that is anesthetic, potentiates GABAergic transmission, and gives an appropriate dissociation constant, Kd Ϸ 0. ferritin ͉ probe ͉ GABA ͉ propofol ͉ imaging

Cited by 47 publications

References 36 publications

A Unitary Anesthetic Binding Site at High Resolution

A Unitary Anesthetic Binding Site at High Resolution

Beyond the detergent effect: a binding site for sodium dodecyl sulfate (SDS) in mammalian apoferritin

H-ferritin–nanocaged doxorubicin nanoparticles specifically target and kill tumors with a single-dose injection

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