Muscarinic toxins from the green mamba

Bradley, Karen N.

doi:10.1016/s0163-7258(99)00064-9

Cited by 63 publications

(49 citation statements)

References 102 publications

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“…This toxin interacts with selectivity at least 20,000 times higher for the M 1 receptor compared with the other subtypes. All the values reported in Table 1 are in good agreement with previous affinities described in the literature (for review, see Bradley, 2000;Karlsson et al, 2000), confirming that the chemically synthesized toxins have pharmacological affinities and specificities (Max et al, 1993;Carsi and Potter, 2000;Olianas et al, 2000) renders the calculation of its true affinity constant difficult, and the use of IC 50 values is preferable. In addition, the large quantity of toxin available by chemical synthesis allowed testing for the first time of the effect of 10 M sMT7 on the M 2 -M 4 receptors, indicating that this toxin interacts preferentially (at least 10,000 times greater affinity) with the M 1 subtype.…”

Section: Discussionsupporting

confidence: 81%

“…Several neurotoxins have been purified from the venom of African mambas (Dendroaspis angusticeps and Dendroaspis polylepis) and characterized for their specific interaction with various muscarinic receptors (for review, see Bradley, 2000;Jerusalinsky et al, 2000;Karlsson et al, 2000;Potter, 2001). These muscarinic toxins are peptides of 63 to 66 residues with four disulfide bonds and a common three-finger fold structure (Segalas et al, 1995).…”

mentioning

confidence: 99%

“…In this context, the selectivity with which muscarinic toxins recognize some muscarinic receptors may be very useful in the identification of the type of receptor expressed in various tissues and could enable their use as potential therapeutic agents in diseases involving muscarinic receptors (Eglen et al, 1999;Bradley, 2000). However, the amount of toxin available from snake venom is often too low to allow their extensive functional characterization.…”

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confidence: 99%

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Chemical Synthesis of MT1 and MT7 Muscarinic Toxins: Critical Role of Arg-34 in Their Interaction with M₁ Muscarinic Receptor

et al. 2003

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Two muscarinic toxins, MT1 and MT7, were obtained by onestep solid-phase synthesis using the 9-fluorenylmethoxycarbonyl-based method. The synthetic and natural toxins, isolated from the snake venom or recombinantly expressed, display identical physicochemical properties and pharmacological profiles. High protein recovery allowed us to specify the selectivity of these toxins for various muscarinic receptor subtypes. Thus, sMT7 has a selectivity for the M 1 receptor that is at least 20,000 times that for the other subtypes. The stability of the toxinreceptor complexes indicates that sMT1 interacts reversibly with the M 1 receptor, unlike sMT7, which binds it quasi-irreversibly. The effect of the synthetic toxins on the atropineinduced [3 H]N-methylscopolamine (NMS) dissociation confirms that sMT7 targets the allosteric site on the M 1 receptor, whereas sMT1 seems interact on the orthosteric one. The great decreases in the binding potencies observed after the R34A modification in sMT1 and sMT7 toxins highlight the functional role of this conserved residue in their interactions with the M 1 receptor. Interestingly, after the R34A modification, the sMT7 toxin binds reversibly on the M 1 receptor. Furthermore, the potency of sMT7-R34A for the NMS-occupied receptor is lower compared with unmodified toxin, supporting the role of this residue in the allosteric interaction of sMT7. All these results and the different charge distributions observed at the two toxin surfaces of their structure models support the hypothesis that the two toxins recognize the M 1 receptor differently.

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Section: Discussionsupporting

confidence: 81%

mentioning

confidence: 99%

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confidence: 99%

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Chemical Synthesis of MT1 and MT7 Muscarinic Toxins: Critical Role of Arg-34 in Their Interaction with M₁ Muscarinic Receptor

et al. 2003

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“…Ten different muscarinic toxins have been isolated so far. Despite their high sequence homology, they show divergent interaction profiles with the different muscarinic receptor subtypes (for review, see Bradley, 2000;Karlsson et al, 2000). For instance, the MT7 toxin, purified from the venom of the green mamba, binds to the M 1 receptor subtype with a potency in the low or subnanomolar range, with at least 10,000-fold selectivity relative to the other receptor subtypes (Max et al, 1993;Carsi and Potter, 2000;Olianas et al, 2000;Mourier et al, 2003).…”

mentioning

confidence: 99%

Different Interactions between MT7 Toxin and the Human Muscarinic M₁ Receptor in Its Free and N-Methylscopolamine-Occupied States

Fruchart-Gaillard

Mourier²,

Marquer³

et al. 2008

Mol Pharmacol

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Muscarinic MT7 toxin is a highly selective and potent antagonist of the M 1 subtype of muscarinic receptor and acts by binding to an allosteric site. To identify the molecular determinants by which MT7 toxin interacts with this receptor in its free and NMS-occupied states, the effect on toxin potency of alanine substitution was evaluated in equilibrium and kinetic binding experiments as well as in functional assays. The determination of the crystallographic structure of an MT7-derivative (MT7-diiodoTyr51) allowed the selection of candidate residues that are accessible and present on both faces of the three toxin loops. The equilibrium binding data are consistent with negative cooperativity between N-methylscopolamine (NMS) and wild-type or modified MT7 and highlight the critical role of the tip of the central loop of the toxin (Arg34, Met35Tyr36) in its interaction with the unoccupied receptor. Examination of the potency of wild-type and modified toxins to allosterically decrease the dissociation rate of [ 3 H]NMS allowed the identification of the MT7 residues involved in its interaction with the NMSoccupied receptor. In contrast to the results with the unoccupied receptor, the most important residue for this interaction was Tyr36 in loop II, assisted by Trp10 in loop I and Arg52 in loop III. The critical role of the tips of the MT7 loops was also confirmed in functional experiments. The high specificity of the MT7-M 1 receptor interaction exploits several MT7-specific residues and reveals a different mode of interaction of the toxin with the free and NMS-occupied states of the receptor.Muscarinic neurotoxins, small peptides of 64 to 66 residues derived from the venom of African mambas (Dendroaspis angusticeps and Dendroaspis polylepis), are well known for their ability to interact with different muscarinic receptor subtypes. NMR and X-ray studies of the MT2 toxin have shown that muscarinic toxins have the three-finger fold structure, characteristic of the large superfamily of toxins that act at cholinergic synapses (Ménez and Ducruix, 1993;Ségalas et al., 1995;Servent and Ménez, 2001). Ten different muscarinic toxins have been isolated so far. Despite their high sequence homology, they show divergent interaction profiles with the different muscarinic receptor subtypes (for review, see Bradley, 2000;Karlsson et al., 2000). For instance, the MT7 toxin, purified from the venom of the green mamba, binds to the M 1 receptor subtype with a potency in the low or subnanomolar range, with at least 10,000-fold selectivity relative to the other receptor subtypes (Max et al., 1993;Carsi and Potter, 2000;Olianas et al., 2000;Mourier et al., 2003 (Olianas et al., 2000(Olianas et al., , 2004Krajewski et al., 2001;Mourier et al., 2003). In summary, MT7 acts as a highly selective antagonist of M 1 receptors by establishing a strong and stable interaction with an allosteric binding site on the receptor.There is a limited understanding of the specificity, selectivity, and mechanism of action of the muscarinic toxins at Ar...

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“…For instance, approximately 10 different muscarinic toxins have been isolated from the venom of African mambas and, despite a high sequence homology, they exhibit clear differences in their functional activities and possess various profiles of interaction with the five muscarinic receptor subtypes (Bradley, 2000;Karlsson et al, 2000;Onali et al, 2005 (Max et al, 1993;Olianas et al, 2000Olianas et al, , 2004Krajewski et al, 2001;Mourier et al, 2003). However, all of these studies were done indirectly, and the mode of action of this toxin with M 1 receptor is not yet fully understood.…”

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confidence: 99%

Identification of Various Allosteric Interaction Sites on M₁Muscarinic Receptor Using¹²⁵I-Met35-Oxidized Muscarinic Toxin 7

Fruchart-Gaillard¹,

Mourier²,

Marquer³

et al. 2006

Mol Pharmacol

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Monoiodinated, Met35-oxidized muscarinic toxin 7 (MT7ox) was synthesized, and its affinity constants for free or N-methyl scopolamine (NMS)-occupied hM 1 receptor were measured directly by equilibrium and kinetic binding experiments. Identical values were obtained with the two types of assay methods, 14 pM and 0.9 nM in free or NMS-liganded receptor states, respectively, highlighting a strong negative cooperativity between this allosteric toxin and NMS. Identical results were obtained with indirect binding experiments with [3 H]NMS using the ternary complex model, clearly demonstrating the reciprocal nature of this cooperativity. Furthermore, the effects of various orthosteric and allosteric agents on the dissociation kinetic of 125 I-MT7ox were measured and show that, except for the MT1 toxin, all of the ligands studied [NMS, atropine, gallamine, brucine, tacrine, staurosporine, and (9S,10S,12R)-2,3,9,10,11-hexahydro-10-hydroxy-9-methyl-1-oxo-9,12-epoxy-1H-diindolo[1,2,3-fg:3Ј,2Ј,1Ј-kl]pyrrolo [3,4-i][1,6]benzodiazocine-10-carboxylic acid hexyl ester (KT5720)] interact allosterically with muscarinic toxin 7. Equilibrium binding experiments with 125 IMT7ox and [3 H]NMS were conducted to reveal the effects of these ligands on the free receptor, and affinity constants (pK x values) were calculated using the allosteric ternary complex model. Our results suggest that MT7 toxin interacts with hM 1 receptor at a specific allosteric site, which may partially overlap those identified previously for "classic" or "atypical" allosteric agents and highlight the potential of this new allosteric tracer in studying allosterism at muscarinic receptors.An increasing body of evidence now shows that many Gprotein-coupled receptors are susceptible to allosteric modulation (Christopoulos and Kenakin, 2002), and this phenomenon is particularly well documented for the five muscarinic acetylcholine receptor subtypes (Tucek and Proska, 1995;Birdsall et al., 1997;Ellis, 1997;Lazareno et al., 2004;Birdsall and Lazareno, 2005). These receptors are characterized by at least two distinct binding sites. Agonists and competitive antagonists bind to the orthosteric site, located inside the transmembrane domain, whereas allosteric agents induce a positive or a negative alteration of the orthosteric ligand affinity by interacting with an allosteric site, located more extracellularly (Stockton et al., 1983;Ellis et al., 1993;Ellis, 1997;Trankle and Mohr, 1997;Christopoulos et al., 1998;Christopoulos and Kenakin, 2002). Furthermore, recent publications report the presence of at least two distinct allosteric sites on muscarinic receptors selective for distinct classes of allosteric agents, suggesting that it is possible for three small ligands to bind simultaneously to a muscarinic receptor (Birdsall et al., 2001;Lazareno et al., 2002). For instance, staurosporine and other indolocarbazole compounds have been shown to interact at a second allosteric site on hM 1 receptor, associated with a positive cooperativity with ACh (Lazareno et al., 20...

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Muscarinic toxins from the green mamba

Cited by 63 publications

References 102 publications

Chemical Synthesis of MT1 and MT7 Muscarinic Toxins: Critical Role of Arg-34 in Their Interaction with M₁ Muscarinic Receptor

Chemical Synthesis of MT1 and MT7 Muscarinic Toxins: Critical Role of Arg-34 in Their Interaction with M₁ Muscarinic Receptor

Different Interactions between MT7 Toxin and the Human Muscarinic M₁ Receptor in Its Free and N-Methylscopolamine-Occupied States

Identification of Various Allosteric Interaction Sites on M₁Muscarinic Receptor Using¹²⁵I-Met35-Oxidized Muscarinic Toxin 7

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Muscarinic toxins from the green mamba

Cited by 63 publications

References 102 publications

Chemical Synthesis of MT1 and MT7 Muscarinic Toxins: Critical Role of Arg-34 in Their Interaction with M1 Muscarinic Receptor

Chemical Synthesis of MT1 and MT7 Muscarinic Toxins: Critical Role of Arg-34 in Their Interaction with M1 Muscarinic Receptor

Different Interactions between MT7 Toxin and the Human Muscarinic M1 Receptor in Its Free and N-Methylscopolamine-Occupied States

Identification of Various Allosteric Interaction Sites on M1Muscarinic Receptor Using125I-Met35-Oxidized Muscarinic Toxin 7

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Chemical Synthesis of MT1 and MT7 Muscarinic Toxins: Critical Role of Arg-34 in Their Interaction with M₁ Muscarinic Receptor

Chemical Synthesis of MT1 and MT7 Muscarinic Toxins: Critical Role of Arg-34 in Their Interaction with M₁ Muscarinic Receptor

Different Interactions between MT7 Toxin and the Human Muscarinic M₁ Receptor in Its Free and N-Methylscopolamine-Occupied States

Identification of Various Allosteric Interaction Sites on M₁Muscarinic Receptor Using¹²⁵I-Met35-Oxidized Muscarinic Toxin 7