Halothane binds in the adenine‐specific niche of crystalline adenylate kinase

Sachsenheimer, W.; Pai, E.F.; Schulz, Georg E.; Schirmer, R. Heiner

doi:10.1016/0014-5793(77)80809-0

Cited by 53 publications

(25 citation statements)

References 20 publications

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“…A blood alcohol level of 0.1%, the legal limit for driving in many US states, corresponds to a concentration of ~22 mM. Structural and biophysical studies of anesthetics bound to model proteins have defined the importance of both non-polar and polar groups in the binding sites of these molecules [19][20][21][22][23] and binding sites for alcohols will also posses such an amphipathic nature 18 .…”

Section: Introductionmentioning

confidence: 99%

Structure of a specific alcohol-binding site defined by the odorant binding protein LUSH from Drosophila melanogaster

Kruse

Zhao

Smith

et al. 2003

Nat Struct Mol Biol

213

265

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SummaryWe have solved the high-resolution crystal structure of the Drosophila melanogaster alcoholbinding protein LUSH in the complexes it forms with a series of short chain n-alcohols. LUSH is the first known non-enzyme protein with a defined in vivo alcohol-binding function. The structure of LUSH reveals a set of molecular interactions that define a specific alcohol-binding site. A group of amino acids, Thr57, Ser52 and Thr48 form a network of concerted hydrogen bonds between the protein and the alcohol that provide a structural motif to increase alcohol binding affinity at this site. This motif appears to be conserved in a number of mammalian ligand-gated ion channels that are directly implicated in the pharmacological effects of alcohol. Further, these sequences are found in regions of ion-channels that are known to infer alcohol sensitivity. We suggest the alcohol-binding site in LUSH represents a general model for alcohol-binding sites in proteins.

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Section: Introductionmentioning

confidence: 99%

Structure of a specific alcohol-binding site defined by the odorant binding protein LUSH from Drosophila melanogaster

Kruse

Zhao

Smith

et al. 2003

Nat Struct Mol Biol

213

265

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“…The Ap5A-induced rotation of the Asp-41-containing nucleotide monophosphate binding domain relative to the His-36-containing CORE domain (28) alters the topography of the anesthetic binding pocket, probably accounting for the decreased photoincorporation of 3-[ 3 H]azioctanol in the presence of Ap5A. A low resolution difference Fourier map of a halothaneadenylate kinase complex (4) showed that halothane (CF 3 ⅐CHClBr) bound to the CORE domain between the central ␤-sheet (strands 1, 4, and 5) and helix ␣1, in an area distant from the alkanol binding site identified in this work. Thus, halothane and alkanols appear to target distinct sites in adenylate kinase.…”

Section: Discussionmentioning

confidence: 99%

“…They have a central domain (CORE) consisting of a five-stranded parallel ␤-sheet surrounded by ␣-helices, an ␣-helical nucleotide monophosphate binding domain, and a region that covers the active site during catalysis (LID) (17,18). Halothane inhibits muscle adenylate kinase (myokinase), an action once implicated in general anestheticinduced malignant hypothermia, and its binding site has been identified (4). At ϳ22 kDa, the small variant adenylate kinases are considerably smaller than the much studied surrogate anesthetic target, luciferase (molecular mass ϳ 72 kDa), allowing us to apply the full power of mass spectrometry.…”

Section: ) 3-[mentioning

confidence: 99%

Geometric Isomers of a Photoactivable General Anesthetic Delineate a Binding Site on Adenylate Kinase

Addona¹,

Husain²,

Stehle³

et al. 2002

Journal of Biological Chemistry

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General anesthetics are a class of drugs whose mode of action is poorly understood. Here, two photoactivable general anesthetics, n-octan-1-ol geometric isomers bearing a diazirine group on either the third or seventh carbon (3-and 7-azioctanol, respectively), were used to locate and delineate an anesthetic site on adenylate kinase. Each photoincorporated at a mole ratio of 1:1 as determined by mass spectrometry. The photolabeled kinase was subjected to tryptic digest, and the fragments were separated by chromatography and sequenced by mass spectrometry. 3-Azioctanol photolabeled His-36, whereas its isomer, 7-azioctanol, photolabeled Asp-41. Inspection of the known structure of adenylate kinase shows that the side chains of these residues are within ϳ5 Å of each other. This distance matches the separation of the 3-and 7-positions of an extended aliphatic chain. The alkanol site so-defined spans two domains of adenylate kinase. His-36 is part of the CORE domain, and Asp-41 belongs to the nucleotide monophosphate binding domain. Upon ligand binding the nucleotide monophosphate binding domain rotates relative to the CORE domain, causing a conformational change that might be expected to affect alkanol binding. Indeed, the substrate-mimicking inhibitor adenosine-(5)-pentaphospho-(5)-adenosine (Ap5A) reduced the photoincorporation of 3-[ 3 H]azioctanol by 75%.

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“…Low resolution x-ray crystallography of adenylate kinase has localized halothane to a discrete interhelical niche lined with hydrophobic residues, as well as more polar aromatics like tyrosine (22). Also shown with x-ray diffraction, the active site of the enzyme haloalkane dehalogenase, which binds and catalyzes the dechlorination of dichloroethane (23), contains two tryptophans and two phenylalanines, whose protons are thought to interact electrostatically with the halogens of the substrate.…”

Section: Discussionmentioning

confidence: 99%

Amino Acid Resolution of Halothane Binding Sites in Serum Albumin

Eckenhoff

1996

Journal of Biological Chemistry

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Saturable binding of various inhaled anesthetics to serum albumin has been shown with a variety of approaches. In order to determine the location of halothane binding sites in serum albumin, both human and bovine serum albumins (HSA and BSA) were photolabeled with [ 14 C]halothane, and subjected to proteolysis and microsequencing. BSA was found to have a higher affinity for halothane than HSA, and it contained two specifically labeled sites. One site was characterized by diffuse labeling from Trp 212 -Leu 217, and the other by a more discrete and higher affinity labeling at Trp 134 -Gly 135. HSA contained only a single labeled site, and although lower affinity, was determined to be analogous to BSA Trp 212 . The position 130 -140 region of HSA, having a leucine instead of tryptophan at position 134, was not labeled. These results demonstrate specific and discrete binding of an inhaled anesthetic to a mammaliansoluble protein, and further suggest the importance of aromatic residues as one feature of inhaled anesthetic binding sites.Volatile inhalational anesthetics can bind to and alter the function of a variety of proteins, both soluble and membranebound (1). Determination of the characteristics of these binding sites in proteins should allow prediction of the interactive forces producing binding (immobilization) and also hint at the structural or dynamical consequences to the target. Furthermore, the demonstration of a binding motif common to many protein targets will simplify the search for anesthetic binding domains in other systems. Unfortunately, the character of anesthetic binding sites in protein has been difficult to determine directly, and few examples are available.While consensus places important actions of anesthetics at membrane protein, the presence of lipid and multiple apparently specific binding sites (2) renders these complex models presently unsuitable for a focused search of binding site features. Certain soluble proteins, on the other hand, appear to have a limited number of discrete binding sites for inhalational anesthetics, making them reasonable initial candidates for characterizing anesthetic binding domains. Accordingly, several approaches have identified specific, saturable binding of halothane in bovine serum albumin (3-6) and have also demonstrated anesthetic-induced alterations in the structure or carrier function (7)(8)(9)(10)(11)(12) C]2-bromo-2-chloro-1,1,1-trifluoroethane; 6.6 mCi/mmol) was purchased from DuPont NEN as the neat compound, which was dissolved in buffer as a stock 6 mM solution and stored at Ϫ80°C. Electrophoresis reagents and supplies were purchased from Bio-Rad.Photoaffinity Labeling-Albumin solutions (generally 15 M) in deoxygenated (argon-bubbled) 150 mM NaCl, containing various concentrations (0.05-0.4 mM) of [ 14 C]halothane was exposed to 254 nm light (Oriel Hg pencil calibration lamp at 7 mm) for 60 -100 s in 5-mm path-length quartz cuvettes (2 ml) at 20 -22°C. The final addition completely filled the cuvette, which was then sealed with a Teflon stopper, ...

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Halothane binds in the adenine‐specific niche of crystalline adenylate kinase

Cited by 53 publications

References 20 publications

Structure of a specific alcohol-binding site defined by the odorant binding protein LUSH from Drosophila melanogaster

Structure of a specific alcohol-binding site defined by the odorant binding protein LUSH from Drosophila melanogaster

Geometric Isomers of a Photoactivable General Anesthetic Delineate a Binding Site on Adenylate Kinase

Amino Acid Resolution of Halothane Binding Sites in Serum Albumin

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