Karlsson E

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Full name : Karlsson Evert

First name : Evert

Mail : Institute of Biochemistry and Organic Chemistry, Uppsala

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Country : Sweden

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References (47)

Title : Weak toxin WTX from Naja kaouthia cobra venom interacts with both nicotinic and muscarinic acetylcholine receptors - Mordvintsev_2009_FEBS.J_276_5065
Author(s) : Mordvintsev DY , Polyak YL , Rodionov DI , Jakubik J , Dolezal V , Karlsson E , Tsetlin VI , Utkin YN
Ref : Febs J , 276 :5065 , 2009
Abstract : Iodinated [125I] weak toxin from Naja kaouthia (WTX) cobra venom was injected into mice, and organ-specific binding was monitored. Relatively high levels of [125I]WTX were detected in the adrenal glands. Rat adrenal membranes were therefore used for analysis of [125I]WTX-binding sites. Specific [125I]WTX binding was partially inhibited by both alpha-cobratoxin, a blocker of the alpha7 and muscle-type nicotinic acetylcholine receptors (nAChRs), and by atropine, an antagonist of the muscarinic acetylcholine receptor (mAChR). Binding to rat adrenal nAChR had a Kd of 2.0+/-0.8 microM and was inhibited by alpha-cobratoxin but not by a short-chain alpha-neurotoxin antagonist of the muscle-type nAChR, suggesting a specific interaction with the alpha7-type nAChR. WTX binding was reduced not only by atropine but also by other muscarinic agents (oxotremorine and muscarinic toxins from Dendroaspis angusticeps), indicating an interaction with mAChR. This interaction was further characterized using individual subtypes of human mAChRs expressed in Chinese hamster ovary cells. WTX concentrations up to 30 microM did not inhibit binding of [3H]acetylcholine to any subtype of mAChR by more than 50%. Depending on receptor subtype, WTX either increased or had no effect on the binding of the muscarinic antagonist [3H]N-methylscopolamine, which binds to the orthosteric site, a finding indicative of an allosteric interaction. Furthermore, WTX alone activated G-protein coupling with all mAChR subtypes and reduced the efficacy of acetylcholine in activating G-proteins with the M1, M4, and M5 subtypes. Our data demonstrate an orthosteric WTX interaction with nAChR and an allosteric interaction with mAChRs.
ESTHER : Mordvintsev_2009_FEBS.J_276_5065
PubMedSearch : Mordvintsev_2009_FEBS.J_276_5065
PubMedID: 19682302

Title : Differential coupling of M1 muscarinic and alpha7 nicotinic receptors to inhibition of pemphigus acantholysis - Chernyavsky_2008_J.Biol.Chem_283_3401
Author(s) : Chernyavsky AI , Arredondo J , Piser T , Karlsson E , Grando SA
Ref : Journal of Biological Chemistry , 283 :3401 , 2008
Abstract : The mechanisms mediating and regulating assembly and disassembly of intercellular junctions is a subject of intensive research. The IgG autoantibodies produced in patients with the immunoblistering skin disease pemphigus vulgaris (PV) can induce keratinocyte (KC) dyshesion (acantholysis) via mechanisms that involve signaling kinases targeting intercellular adhesion molecules, thus providing a useful model to study the physiologic regulation of KC cohesion. Previous studies showed that activation of Src and protein kinase C are the earliest events in the PV IgG-induced intracellular phosphorylation cascades and that cholinergic agonists are effective for treating patients with pemphigus. In this study, we sought to elucidate the molecular mechanisms allowing cholinergic agonists to inhibit PV IgG-induced acantholysis and phosphorylation of KC adhesion molecules. The extent of acantholysis in KC monolayers correlated closely with the degree of PV IgG-induced phosphorylation of p120- and beta-catenins, with classic isoforms of protein kinase C mediating serine phosphorylation of beta-catenin and Src-tyrosine phosphorylation of p120-catenin. The M(1) muscarinic agonist pilocarpine blocked phosphorylation of both catenins, which could be abolised by the M(1) antagonist MT7. The alpha7 nicotinic agonist AR-R17779 inhibited phosphorylation of P120-cateinin. The alpha7 antagonist methyllycaconitine abolished the effect of AR-R17779. Okadaic acid abrogated protective effects of agonists on phosphorylation of beta-catenin, and pervanadate, on that of p120-catenin. Stimulation of KCs with pilocarpine significantly (p < 0.05) elevated both serine/threonine and tyrosine phosphatase activities in KCs. AR-R17779 both stimulated tyrosine phosphatase and decreased PV IgG-induced Src activity. Methyllycaconitine released Src activity in intact KCs and caused acantholysis. Thus, downstream signaling from M(1) abolished PV IgG-dependent catenin phosphorylation due to activation of both serine/threonine and tyrosine phosphatases, whereas alpha7 action involved both activation of tyrosine phosphatase and inhibition of Src. These findings identified novel paradigm of regulation of signaling kinases associated with cholinergic receptors and provided mechanistic explanation of therapeutic activity of cholinomimetics in PV patients.
ESTHER : Chernyavsky_2008_J.Biol.Chem_283_3401
PubMedSearch : Chernyavsky_2008_J.Biol.Chem_283_3401
PubMedID: 18073210

Title : Temporal and region-dependent changes in muscarinic M4 receptors in the hippocampus and entorhinal cortex of adrenalectomized rats - Mulugeta_2006_Exp.Brain.Res_173_309
Author(s) : Mulugeta E , Chandranath I , Karlsson E , Winblad B , Adem A
Ref : Experimental Brain Research , 173 :309 , 2006
Abstract : Long-term adrenalectomy induces a dramatic loss of cells in the dentate gyrus and CA1-CA4 fields of the hippocampus resulting in an impairment of cognitive functions such as spatial learning, memory and exploratory behaviour. Muscarinic M1 and M4 receptor levels in the hippocampus and entorhinal cortex of adult male Wistar rats were examined 3, 14, 30, 90, and 150 days after adrenalectomy. Receptor levels in the entorhinal cortex and the hippocampus were determined by quantitative autoradiography using 125I-M1-toxin-1 and 125I-M4-toxin-1, M1 and M4 subtype selective antagonists, respectively. Moreover, the level of hippocampal M1 and M4 muscarinic receptors were evaluated 1 month after adrenalectomy by immunoblot analysis. Adrenalectomy induced apoptotic processes were examined by analysing apoptotic markers using Western blot analysis. No significant changes were observed in the level of muscarinic M1 receptors in the entorhinal cortex, the dentate gyrus and in the different CA fields of the hippocampus of adrenalectomized (ADX) rats. However, M4 receptors showed a significant decrease in the entorhinal cortex (at 3 days), dentate gyrus and CA4 (at 14 days), CA3 (at 30 days), and CA2 and CA1 (at 90 days) after adrenalectomy. Moreover, a decrease in the level of M4 receptors was detected in ADX rats 1 month after adrenalectomy as compared with sham groups using M4 specific antibody. Apoptotic markers such as PARP and p53 were significantly increased whereas Bcl-2 marker was decreased in ADX rat brain homogenates compared to controls. Our results show that M1 and M4 receptors are differentially affected by adrenalectomy and indicate that these subtypes have different functions in the hippocampus. Our data on time and region-dependent decreases in hippocampal M4 receptors indicate that the M4 receptor subtype is influenced by adrenal hormones and suggest that the M4 receptor might be linked to memory function in the hippocampus.
ESTHER : Mulugeta_2006_Exp.Brain.Res_173_309
PubMedSearch : Mulugeta_2006_Exp.Brain.Res_173_309
PubMedID: 16676164

Title : The Ras\/Raf-1\/MEK1\/ERK signaling pathway coupled to integrin expression mediates cholinergic regulation of keratinocyte directional migration - Chernyavsky_2005_J.Biol.Chem_280_39220
Author(s) : Chernyavsky AI , Arredondo J , Karlsson E , Wessler I , Grando SA
Ref : Journal of Biological Chemistry , 280 :39220 , 2005
Abstract : The physiologic mechanisms that determine directionality of lateral migration are a subject of intense research. Galvanotropism in a direct current (DC) electric field represents a natural model of cell re-orientation toward the direction of future migration. Keratinocyte migration is regulated through both the nicotinic and muscarinic classes of acetylcholine (ACh) receptors. We sought to identify the signaling pathway mediating the cholinergic regulation of chemotaxis and galvanotropism. The pharmacologic and molecular modifiers of the Ras/Raf-1/MEK1/ERK signaling pathway altered both chemotaxis toward choline and galvanotropism toward the cathode in a similar way, indicating that the same signaling steps were involved. The galvanotropism was abrogated due to inhibition of ACh production by hemicholinium-3 and restored by exogenously added carbachol. The concentration gradients of ACh and choline toward the cathode in a DC field were established by high-performance liquid chromatographic measurements. This suggested that keratinocyte galvanotaxis is, in effect, chemotaxis toward the concentration gradient of ACh, which it creates in a DC field due to its highly positive charge. A time-course immunofluorescence study of the membrane redistribution of ACh receptors in keratinocytes exposed to a DC field revealed rapid relocation to and clustering at the leading edge of alpha7 nicotinic and M(1) muscarinic receptors. Their inactivation with selective antagonists or small interfering RNAs inhibited galvanotropism, which could be prevented by transfecting the cells with constitutively active MEK1. The end-point effect of the cooperative signaling downstream from alpha7 and M(1) through the MEK1/ERK was an up-regulated expression of alpha(2) and alpha(3) integrins, as judged from the results of real-time PCR and quantitative immunoblotting. Thus, alpha7 works together with M(1) to orient a keratinocyte toward direction of its future migration. Both alpha7 and M(1) apparently engage the Ras/Raf/MEK/ERK pathway to up-regulate expression of the "sedentary" integrins required for stabilization of the lamellipodium at the keratinocyte leading edge.
ESTHER : Chernyavsky_2005_J.Biol.Chem_280_39220
PubMedSearch : Chernyavsky_2005_J.Biol.Chem_280_39220
PubMedID: 16150734

Title : The pharmacological action of MT-7 - Onali_2005_Life.Sci_76_1547
Author(s) : Onali P , Adem A , Karlsson E , Olianas MC
Ref : Life Sciences , 76 :1547 , 2005
Abstract : The mamba toxin MT-7 is the most selective ligand currently available for the muscarinic M1 receptor subtype. The toxin binds stably to the receptor and blocks the agonist-induced activation non-competitively. Although its mode of action on M1 receptors is not yet fully understood, some of the toxin properties support an allosteric mechanism. Thus, the toxin fails to elicit a complete inhibition of the binding of either the muscarinic antagonist [3H]N-methyl-scopolamine ([3H]NMS) or the agonist [3H]acetylcholine ([3H]ACh). When added to ligand-occupied M1 receptors, the toxin slows the dissociation rate of [3H]NMS and increases that of [3H]ACh. Site-directed mutagenesis studies have provided important information about the toxin amino acid residues which are critical for the stable binding to the receptor and for the allosteric modulation of antagonist dissociation. In vivo studies have shown that the intracerebral injection of MT-7 causes a long-lasting blockade of M1 receptor, thus providing a tool for the characterization of the functional role of this receptor subtype in discrete brain areas.
ESTHER : Onali_2005_Life.Sci_76_1547
PubMedSearch : Onali_2005_Life.Sci_76_1547
PubMedID: 15680165

Title : Novel signaling pathways mediating reciprocal control of keratinocyte migration and wound epithelialization through M3 and M4 muscarinic receptors - Chernyavsky_2004_J.Cell.Biol_166_261
Author(s) : Chernyavsky AI , Arredondo J , Wess J , Karlsson E , Grando SA
Ref : Journal of Cell Biology , 166 :261 , 2004
Abstract : To test the hypothesis that keratinocyte (KC) migration is modulated by distinct muscarinic acetylcholine (ACh) receptor subtypes, we inactivated signaling through specific receptors in in vitro and in vivo models of reepithelialization by subtype-selective antagonists, small interfering RNA, and gene knockout in mice. KC migration and wound reepithelialization were facilitated by M4 and inhibited by M3. Additional studies showed that M4 increases expression of "migratory" integrins alpha5beta1, alphaVbeta5, and alphaVbeta6, whereas M3 up-regulates "sedentary" integrins alpha2beta1 and alpha3beta1. Inhibition of migration by M3 was mediated through Ca2+-dependent guanylyl cyclase-cyclic GMP-protein kinase G signaling pathway. The M4 effects resulted from inhibition of the inhibitory pathway involving the adenylyl cyclase-cyclic AMP-protein kinase A pathway. Both signaling pathways intersected at Rho, indicating that Rho kinase provides a common effector for M3 and M4 regulation of cell migration. These findings offer novel insights into the mechanisms of ACh-mediated modulation of KC migration and wound reepithelialization, and may aid the development of novel methods to promote wound healing.
ESTHER : Chernyavsky_2004_J.Cell.Biol_166_261
PubMedSearch : Chernyavsky_2004_J.Cell.Biol_166_261
PubMedID: 15263021

Title : Action of the muscarinic toxin MT7 on agonist-bound muscarinic M1 receptors - Olianas_2004_Eur.J.Pharmacol_487_65
Author(s) : Olianas MC , Adem A , Karlsson E , Onali P
Ref : European Journal of Pharmacology , 487 :65 , 2004
Abstract : The muscarinic toxin MT7 is the most selective ligand for the muscarinic M(1) receptors. Previous studies have shown that the toxin interacts with the antagonist-receptor complex and slows the antagonist dissociation rate, possibly by binding to an allosteric site and impeding the access to and egress from the orthosteric binding pocket. In the present study, we investigated the action of MT7 on agonist-occupied receptors in functional and radioligand binding assays of the cloned human muscarinic M(1) receptor expressed in Chinese hamster ovary cells. In time-course experiments, the addition of MT7 rapidly blocked the acetylcholine-stimulated guanosine-5'-O-(3-[(35)S]thio)triphosphate binding to membrane G proteins. Similarly, in acetylcholine-treated cells MT7 completely stopped the agonist-stimulated [(3)H]inositol phosphate accumulation. In dissociation experiments using membranes pre-equilibrated with [(3)H]acetylcholine, the addition of MT7 increased the rate of radioligand dissociation. The data indicate that MT7, while partially stabilizing the antagonist-receptor complex, effectively destabilizes the agonist-occupied muscarinic M(1) receptors.
ESTHER : Olianas_2004_Eur.J.Pharmacol_487_65
PubMedSearch : Olianas_2004_Eur.J.Pharmacol_487_65
PubMedID: 15033377

Title : Loss of muscarinic M4 receptors in spinal cord of arthritic rats: implications for a role of M4 receptors in pain response - Mulugeta_2003_Brain.Res_982_284
Author(s) : Mulugeta E , El-Bakri N , Karlsson E , Elhassan A , Adem A
Ref : Brain Research , 982 :284 , 2003
Abstract : Changes in the levels of muscarinic M4 receptors in spinal cord of acute and chronic arthritic rats (animal models of pain) were studied by receptor autoradiography using muscarinic M4 receptor subtype selective ligand. Arthritis was induced in female Lewis rats by single intradermal injection of heat-killed Mycobacterium butyricum and sacrificed 12 days (acute group) and 30 days (chronic and control groups) after induction of arthritis. Our results demonstrate significant reduction in the level of M4 receptors in the spinal cord (Rexed laminae I-X) of acute and chronic arthritic rats compared to controls. These findings suggest that the muscarinic M4 receptor subtype may be involved in cholinergic mechanisms of analgesia.
ESTHER : Mulugeta_2003_Brain.Res_982_284
PubMedSearch : Mulugeta_2003_Brain.Res_982_284
PubMedID: 12915263

Title : Loss of muscarinic M4 receptors in hippocampus of Alzheimer patients - Mulugeta_2003_Brain.Res_960_259
Author(s) : Mulugeta E , Karlsson E , Islam A , Kalaria R , Mangat H , Winblad B , Adem A
Ref : Brain Research , 960 :259 , 2003
Abstract : We assessed muscarinic M(1), M(2) and M(4) receptor subtypes in the hippocampus of Alzheimer's and control brains by receptor autoradiography using ligands such as [(125)I]muscarinic toxin-1 ([(125)I]MT-1, M(1) selective), [(3)H]AFDX-384 (M(2) partially selective) and [(125)I]muscarinic toxin 4 ([(125)I]M(4) toxin-1, M(4) selective). Our results revealed a significant decrease in muscarinic M(4) receptor binding in the dentate gyrus and CA4 regions of brain sections from Alzheimer's patients compared to controls. No changes in the density of M(1) or M(2) receptor binding were observed. Our findings suggest that, relative to other muscarinic receptor subtypes, the M(4) receptor could be the subtype which is selectively compromised in Alzeheimer's disease (AD).
ESTHER : Mulugeta_2003_Brain.Res_960_259
PubMedSearch : Mulugeta_2003_Brain.Res_960_259
PubMedID: 12505680

Title : Regulation of acetylcholine release by muscarinic receptors at the mouse neuromuscular junction depends on the activity of acetylcholinesterase - Minic_2002_Eur.J.Neurosci_15_439
Author(s) : Minic J , Molgo J , Karlsson E , Krejci E
Ref : European Journal of Neuroscience , 15 :439 , 2002
Abstract : Muscarinic acetylcholine receptors (mAChRs) play an important role in regulating the release of acetylcholine (ACh) in various tissues. We used subtype-specific antibodies and a fluorescent-labelled muscarinic toxin to demonstrate that mammalian neuromuscular junction expresses mAChR subtypes M1 to M4, and that localization of all subtypes is highly restricted to the innervated part of the muscle. To elucidate the roles of the mAChR subtypes regulating ACh release, we measured the mean quantal content of endplate potentials in isolated mouse phrenic--hemidiaphragm preparations in which release was reduced by a low Ca2+/high Mg2+ medium. Muscarine decreased evoked ACh release in normal junctions but, depending on the concentration, reduced or increased transmitter release in collagen Q-deficient junctions completely lacking acetylcholinesterase (AChE). Both effects were also seen in normal junctions when AChE was inhibited by various doses of fasciculin-2. Block of mAChRs by atropine had no effect on evoked release at normal junctions, but decreased release at junctions lacking AChE. The muscarine-elicited depression of ACh release in normal junctions was completely abolished by pertussis toxin or methoctramine pretreatment, but was not affected by muscarinic toxin MT-3, thus indicating the involvement of the M2 mAChR. The muscarine-induced increase of ACh release in AChE-deficient junctions was not affected by pertussis toxin, but was completely blocked by MT-7, a specific M1 mAChR antagonist. Our results show that the M1 and M2 mAChRs have opposite presynaptic functions in modulating quantal ACh release, and that regulation of release by the two receptor subtypes depends on the functional state of AChE at the neuromuscular junction.
ESTHER : Minic_2002_Eur.J.Neurosci_15_439
PubMedSearch : Minic_2002_Eur.J.Neurosci_15_439
PubMedID: 11876771

Title : Estrogen and progesterone treatment: effects on muscarinic M(4) receptor subtype in the rat brain - El-Bakri_2002_Brain.Res_948_131
Author(s) : El-Bakri NK , Adem A , Suliman IA , Mulugeta E , Karlsson E , Lindgren JU , Winblad B , Islam A
Ref : Brain Research , 948 :131 , 2002
Abstract : We investigated the effect of ovariectomy (OVX) and hormonal treatment for 10 weeks by estradiol and progesterone on muscarinic M(4) receptor subtype in different brain areas of female rats. Moreover, motor activity of OVX and hormone-treated rats was measured by automated open field exploration boxes. Receptor quantification in the hippocampus, frontal cortex, parietal cortex, amygdala and hypothalamus was done by receptor autoradiography using a selective ligand for muscarinic M(4) receptors. Ovariectomy up-regulated M(4) receptors in the dentate gyrus, CA1, CA3, frontal cortex and hypothalamus whereas the estrogen treatment restored M(4) binding to that of the sham group. Progesterone treatment had no effect on the ovariectomy-induced up-regulation of M(4) receptors. Ovariectomy significantly decreased the exploratory activity of the rats compared to the sham group. Estrogen treatment restored the exploratory behavior of the ovariectomized rats to that of the sham group whereas the progesterone-treated rats were less alert to the surrounding when compared to the sham and estrogen supplemented rats. The effect of estrogen on the hippocampal muscarinic M(4) receptor subtype is a novel finding and may have functional significance for cholinergic receptors especially in relation to postmenopausal memory problems and neurodegenerative disease like Alzheimer's disease.
ESTHER : El-Bakri_2002_Brain.Res_948_131
PubMedSearch : El-Bakri_2002_Brain.Res_948_131
PubMedID: 12383964

Title : Kinetic evidence for different mechanisms of interaction of black mamba toxins MT alpha and MT beta with muscarinic receptors - Jolkkonen_2001_Toxicon_39_377
Author(s) : Jolkkonen M , Oras A , Toomela T , Karlsson E , Jarv J , Akerman KE
Ref : Toxicon , 39 :377 , 2001
Abstract : By studying the influence of two toxins from the black mamba Dendroaspis polylepis on the kinetics of [3H]-N-methylscopolamine binding to muscarinic acetylcholine receptors from rat cerebral cortex, it was revealed that these toxins, MT alpha and MT beta, interact with the receptors via kinetically distinct mechanisms. MT beta bound to receptors in a one-step, readily reversible process with the dissociation constant K(d)=5.3 microM. The binding mechanism of MTalpha was more complex, involving at least two consecutive steps. A fast receptor-toxin complex formation (K(T)=3.8 microM) was followed by a slow process of isomerisation of this complex (k(i)=1.8 x 10(-2) s(-1), half-time 39 s). A similar two-step interaction mechanism has been established for a related toxin, MT2 from the green mamba D. angusticeps (K(T)=1.4 microM, k(i)=8.3 x 10(-4) s(-1), half-time 840 s). The slow isomerisation process delays the effect of MT alpha and MT2, but increases their apparent potency compared to toxins unable to induce the isomerisation process.
ESTHER : Jolkkonen_2001_Toxicon_39_377
PubMedSearch : Jolkkonen_2001_Toxicon_39_377
PubMedID: 10978757

Title : Snake toxins with high selectivity for subtypes of muscarinic acetylcholine receptors - Karlsson_2000_Biochimie_82_793
Author(s) : Karlsson E , Jolkkonen M , Mulugeta E , Onali P , Adem A
Ref : Biochimie , 82 :793 , 2000
Abstract : There are five subtypes of muscarinic acetylcholine receptors (M(1) to M(5)) which control a large number of physiological processes, such as the function of heart and smooth muscles, glandular secretion, release of neurotransmitters, gene expression and cognitive functions as learning and memory. A selective ligand is very useful for studying the function of a subtype in presence of other subtypes, which is the most common situation, since a cell or an organ usually has several subtypes. There are many non-selective muscarinic ligands, but only few selective ones. Mambas, African snakes of genus Dendroaspis have toxins, muscarinic toxins, that are selective for M(1), M(2) and M(4) receptors. They consist of 63-66 amino acids and four disulfides which form four loops. They are members of a large group of snake toxins, three-finger toxins; three loops are extended like the middle fingers of a hand and the disulfides and the shortest loop are in the palm of the hand. Some of the toxins target the allosteric site which is located in a cleft of the receptor molecule close to its extracellular part. A possible explanation to the good selectivity is that the toxins bind to the allosteric site, but because of their size they probably also bind to extracellular parts of the receptors which are rather different in the various subtypes. Some other allosteric ligands also have good selectivity, the alkaloid brucine and derivatives are selective for M(1), M(3) and M(4) receptors. Muscarinic toxins have been used in several types of experiments. For instance radioactively labeled M(1) and M(4) selective toxins were used in autoradiography of hippocampus from Alzheimer patients. One significant change in the receptor content was detected in one region of the hippocampus, dentate gyrus, where M(4) receptors were reduced by 50% in patients as compared to age-matched controls. Hippocampus is essential for memory consolidation. M(4) receptors in dentate gyrus may play a role, since they decreased in Alzheimers disease which destroys the memory. Another indication of the role of M(4) receptors for memory is that injection of the M(4) selective antagonist muscarinic toxin 3 (M(4)-toxin 1) into rat hippocampus produced amnesia.
ESTHER : Karlsson_2000_Biochimie_82_793
PubMedSearch : Karlsson_2000_Biochimie_82_793
PubMedID: 11086210

Title : Inhibition of acetylcholine muscarinic M(1) receptor function by the M(1)-selective ligand muscarinic toxin 7 (MT-7) - Olianas_2000_Br.J.Pharmacol_131_447
Author(s) : Olianas MC , Maullu C , Adem A , Mulugeta E , Karlsson E , Onali P
Ref : British Journal of Pharmacology , 131 :447 , 2000
Abstract : MT-7 (1 - 30 nM), a peptide toxin isolated from the venom of the green mamba Dendroaspis angusticeps and previously found to bind selectively to the muscarinic M(1) receptor, inhibited the acetylcholine (ACh)-stimulated [(35)S]-guanosine-5'-O-(3-thio)triphosphate ([(35)S]-GTPgammaS) binding to membranes of Chinese hamster ovary (CHO) cells stably expressing the cloned human muscarinic M(1) receptor subtype. MT-7 failed to affect the ACh-stimulated [(35)S]-GTPgammaS binding in membranes of CHO cells expressing either the M(2), M(3) or M(4) receptor subtype. In N1E-115 neuroblastoma cells endogenously expressing the M(1) and M(4) receptor subtypes, MT-7 (0.3 - 3.0 nM) inhibited the carbachol (CCh)-stimulated inositol phosphates accumulation, but failed to affect the CCh-induced inhibition of pituitary adenylate cyclase activating polypeptide (PACAP) 38-stimulated cyclic AMP accumulation. In both CHO/M(1) and N1E-115 cells the MT-7 inhibition consisted in a decrease of the maximal agonist effect with minimal changes in the agonist EC(50) value. In CHO/M(1) cell membranes, MT-7 (0.05 - 25 nM) reduced the specific binding of 0.05, 1.0 and 15 nM [(3)H]-N-methylscopolamine ([(3)H]-NMS) in a concentration-dependent manner, but failed to cause a complete displacement of the radioligand. Moreover, MT-7 (3 nM) decreased the dissociation rate of [(3)H]-NMS by about 5 fold. CHO/M(1) cell membranes preincubated with MT-7 (10 nM) and washed by centrifugation and resuspension did not recover control [(3)H]-NMS binding for at least 8 h at 30 degrees C. It is concluded that MT-7 acts as a selective noncompetitive antagonist of the muscarinic M(1) receptors by binding stably to an allosteric site.
ESTHER : Olianas_2000_Br.J.Pharmacol_131_447
PubMedSearch : Olianas_2000_Br.J.Pharmacol_131_447
PubMedID: 11015294

Title : Recombinant expression of a selective blocker of M(1) muscarinic receptors - Nasman_2000_Biochem.Biophys.Res.Commun_271_435
Author(s) : Nasman J , Jolkkonen M , Ammoun S , Karlsson E , Akerman KE
Ref : Biochemical & Biophysical Research Communications , 271 :435 , 2000
Abstract : Mamba venoms contain peptides with high selectivity for muscarinic receptors. Due to the limited availability of the M(1) muscarinic receptor-selective MT7 or m1-toxin 1, the peptide was expressed in Sf9 cells using a synthetic cDNA and purified. The isolated peptide had over four orders of magnitude higher affinity for the M(1) compared to M(2)-M(5) muscarinic receptors. The peptide strongly inhibited Ca(2+) mobilisation through recombinant and endogenously expressed M(1) receptors, having no effect on the function of the other subtypes. The MT7 peptide provides a unique tool for identification and functional characterisation of M(1) receptors in cells and tissues.
ESTHER : Nasman_2000_Biochem.Biophys.Res.Commun_271_435
PubMedSearch : Nasman_2000_Biochem.Biophys.Res.Commun_271_435
PubMedID: 10799315

Title : Selectivity profile of muscarinic toxin 3 in functional assays of cloned and native receptors - Olianas_1999_J.Pharmacol.Exp.Ther_288_164
Author(s) : Olianas MC , Ingianni A , Maullu C , Adem A , Karlsson E , Onali P
Ref : Journal of Pharmacology & Experimental Therapeutics , 288 :164 , 1999
Abstract : By using acetylcholine-induced stimulation of [35S]guanosine-5'-O-(3-thio)triphosphate ([35S]GTPgammaS) binding to membrane G proteins as a functional assay of the cloned human m1-m4 muscarinic receptor subtypes stably expressed in Chinese hamster ovary cells, muscarinic toxin 3 (MT3) was found to block the m4 receptor with a potency (pA2 = 8.33) much higher than those displayed at the m1 (pA2 = 6.78), m3 (pA2 = 6.3), and m2 (pA2 < 6.3) subtypes. In N1E-115 cells, which have been reported to express m4 receptors coupled to inhibition of cAMP, MT3 potently antagonized the carbachol-induced inhibition of adenylyl cyclase with a pA2 of 8. 81 and displayed monophasic inhibitory curves. Unexpectedly, in NG108-15 cells, known to express only m4 receptors, MT3 counteracted the carbachol inhibition of adenylyl cyclase with a lower potency (pA2 = 7.60) and showed a biphasic inhibitory curve, suggesting the participation of both m4 and m2 receptors. This possibility was supported by radioligand binding data showing that MT3 failed to completely displace the binding of [3H]N-methylscopolamine to NG108-15 cell membranes and by reverse transcription-polymerase chain reaction analysis, revealing the presence of mRNAs for both m4 and m2 receptor subtypes. These data demonstrate that MT3 possesses a high functional receptor selectivity for both the cloned and native m4 receptors and that in cell systems containing m4 and m2 receptors coupled to a common response, the toxin constitutes a powerful tool to resolve the relative contribution by each receptor subtype.
ESTHER : Olianas_1999_J.Pharmacol.Exp.Ther_288_164
PubMedSearch : Olianas_1999_J.Pharmacol.Exp.Ther_288_164
PubMedID: 9862767

Title : Poster: Detection of muscarinic receptor subtypes in human heart using a reverse transcriptase-polymerase chain reaction (RT-PCR) -
Author(s) : Adem A , Blange I , Sylven C , Winblad E , Mulugeta E , Karlsson E , Mustafa A
Ref : Life Sciences , 64 :591 , 1999
PubMedID:

Title : Identification of rat brain muscarinic M4 receptors coupled to cyclic AMP using the selective antagonist muscarinic toxin 3 - Olianas_1998_Eur.J.Pharmacol_357_235
Author(s) : Olianas MC , Adem A , Karlsson E , Onali P
Ref : European Journal of Pharmacology , 357 :235 , 1998
Abstract : In membranes of olfactory tubercle and striatum, the selective muscarinic M4 receptor antagonist muscarinic toxin 3 completely antagonized the acetylcholine-induced inhibition of forskolin- and dopamine D1 receptor-stimulated cyclic AMP formation with Ki values of 7 and 4 nM, respectively. In olfactory bulb, where acetylcholine stimulated basal adenylyl cyclase activity and inhibited forskolin-stimulated enzyme activity, muscarinic toxin 3 caused a partial antagonism of both acetylcholine effects with high potencies (Ki values = 4-6 nM). In frontal cortex, muscarinic toxin 3 counteracted the acetylcholine-induced potentiation of corticotropin-releasing hormone-stimulated cyclic AMP with a Ki of 58 nM, which is close to the toxin affinity for the muscarinic M1 receptor. In the same brain region, the acetylcholine inhibition of forskolin-stimulated enzyme activity was not affected by muscarinic toxin 3. In microdissected regions of the hippocampus, a significant portion (33-48%) of the acetylcholine inhibition of forskolin-stimulated adenylyl cyclase activity was blocked by muscarinic toxin 3 with Ki values (6-8 nM) consistent with the involvement of muscarinic M4 receptors. These data show that muscarinic toxin 3 discriminates between adenylyl cyclase-coupled muscarinic receptors and demonstrate the utility of the toxin in identifying the relative contribution by the muscarinic M4 receptor subtype.
ESTHER : Olianas_1998_Eur.J.Pharmacol_357_235
PubMedSearch : Olianas_1998_Eur.J.Pharmacol_357_235
PubMedID: 9797042

Title : Localization of M1 muscarinic receptors in rat brain using selective muscarinic toxin-1 - Adem_1997_Brain.Res.Bull_44_597
Author(s) : Adem A , Jolkkonen M , Bogdanovic N , Islam A , Karlsson E
Ref : Brain Research Bulletin , 44 :597 , 1997
Abstract : Mambas, African snakes of the genus Dendroaspis, produce several types of toxins that are of pharmacological interest. The novel muscarinic toxin-1 (MT-1), from the green mamba Dendroaspis angusticeps, binds specifically to muscarinic M1 receptors in homogenates of rat cerebral cortex. Iodination of the toxin, 125I-muscarinic toxin-1 (125I-MT-1), renders the toxin selective for M1 muscarinic receptors. Quantitative measurement of 125I-MT-1 autoradiography in rat brain sections indicated highest labeling in the nucleus accumbens, striatum, and dentate gyrus. High densities of 125I-MT-1 binding sites were located in the CA1 region of the hippocampus, frontal, and parietal cortices. Moderate densities of binding sites were seen in temporal cortex, and hippocampal subregions CA2, CA3, and CA4, whereas low labeling was observed in the cerebellum and spinal cord.
ESTHER : Adem_1997_Brain.Res.Bull_44_597
PubMedSearch : Adem_1997_Brain.Res.Bull_44_597
PubMedID: 9365803

Title : Muscarinic receptor subtype selective toxins - Adem_1997_Life.Sci_60(13-14)_1069
Author(s) : Adem A , Karlsson E
Ref : Life Sciences , 60 :1069 , 1997
Abstract : The muscarinic acetylcholine receptors are monomeric proteins with seven hydrophobic, membrane spanning helices, and share a common evolutionary origin with the other members of the superfamily of membrane proteins known as seven-helix receptors. The amino acid sequences of five different muscarinic acetylcholine receptors, called m1, m2, m3, m4 and m5 have been determined. The five subtypes are expressed to different extent in different tissues. A large number of low molecular ligands for muscarinic receptors are known, but they bind to all five subtypes of receptors and only a few of them have a slightly higher (five-six fold) affinity for one of the subtypes, e.g. pirenzepine for M1 (1) and tripitramine for M2 receptors (2). Several neurotoxins have been isolated from snake venoms and used as pharmacological tools. Mambas, African snakes of genus Dendroaspis, have toxins that recognize muscrinic receptors and some of these muscarinic toxins are the most selective ligands for M1 and M4 receptors known to date.
ESTHER : Adem_1997_Life.Sci_60(13-14)_1069
PubMedSearch : Adem_1997_Life.Sci_60(13-14)_1069
PubMedID: 9121349

Title : Workshop: the use of muscarinic toxins in the study of muscarinic receptors - Jerusalinsky_1997_Life.Sci_60(13-14)_1161
Author(s) : Jerusalinsky D , Harvey A , Karlsson E , Potter LT
Ref : Life Sciences , 60 :1161 , 1997
Abstract : One of the most interesting recent developments in the pharmacology of muscarinic receptors has been the finding of small proteins in the venoms of mamba snakes that bind with high affinity and selectivity to different subtypes of muscarinic receptors. In the workshop on muscarinic toxins, the practicalities of isolating, characterising and using these toxins as tools in the study of muscarinic receptors were discussed.
ESTHER : Jerusalinsky_1997_Life.Sci_60(13-14)_1161
PubMedSearch : Jerusalinsky_1997_Life.Sci_60(13-14)_1161
PubMedID: 9121361

Title : Rat striatal muscarinic receptors coupled to the inhibition of adenylyl cyclase activity: potent block by the selective m4 ligand muscarinic toxin 3 (MT3) - Olianas_1996_Br.J.Pharmacol_118_283
Author(s) : Olianas MC , Adem A , Karlsson E , Onali P
Ref : British Journal of Pharmacology , 118 :283 , 1996
Abstract : 1. In rat striatal membranes, muscarinic toxin 3 (MT3), a selective ligand of the cloned m4 receptor subtype, antagonized the acetylcholine (ACh) inhibition of forskolin- and dopamine D1 receptor-stimulated adenylyl cyclase activities with pA2 values of 8.09 and 8.15, respectively. 2. In radioligand binding experiments, MT3 increased the Kd but did not change the Bmax value of [3H]-N-methylscopolamine (3H]-NMS) binding to rat striatal muscarinic receptors. The toxin displaced the major portion of the [3H]-NMS binding sites with a Ki of 8.0 nM. 3. In rat myocardium, MT3 antagonized the ACh inhibition of adenylyl cyclase with a Ki value of 860 nM. 4. In rat cerebral cortical membranes prelabelled with [3H]-myo-inositol, MT3 counteracted the methacholine stimulation of [3H]-inositol phosphates formation with a Ki value of 113 nM. 5. The present study shows that MT3 is a potent antagonist of the striatal muscarinic receptors coupled to inhibition of adenylyl cyclase activity. This finding provides strong evidence for the classification of these receptors as pharmacologically equivalent to the m4 gene product (M4). On the other hand, the weaker potencies of MT3 in antagonizing the muscarinic responses in cerebral cortex and in the heart are consistent with the reported lower affinities of the toxin for the cloned m1 and m2 receptor subtypes, respectively.
ESTHER : Olianas_1996_Br.J.Pharmacol_118_283
PubMedSearch : Olianas_1996_Br.J.Pharmacol_118_283
PubMedID: 8735628

Title : Fasciculin: modification of carboxyl groups and discussion of structure-activity relationship - Cervenansky_1996_Toxicon_34_718
Author(s) : Cervenansky C , Duran R , Karlsson E
Ref : Toxicon , 34 :718 , 1996
Abstract : Norleucine methylester was coupled to carboxylates of fasciculin 2, a snake toxin that inhibits acetylcholinesterase (AChE). This neutralized negative charges but had no effect on the activity, suggesting that carboxyls do not participate in binding to AChE. Earlier results are discussed. Modification of three aromatic amino acids in the peripheral site of AChE, the binding site for fasciculin, decreased the affinity 100 to one million times. Neutralizing the charge of cationic groups of fasciculin lowered the affinity only three to seven times. A change in either the toxin or enzyme part of a binding site should have about the same effect. Since this was not so, it suggests that cationic groups of fasciculin do not bind to aromatic rings in the peripheral site.
ESTHER : Cervenansky_1996_Toxicon_34_718
PubMedSearch : Cervenansky_1996_Toxicon_34_718
PubMedID: 8817817

Title : Effect of fasciculin on hydrolysis of neutral and choline esters by butyrylcholinesterase, cobra venom and chicken acetylcholinesterases - Duran_1996_Toxicon_34_959
Author(s) : Duran R , Cervenansky C , Karlsson E
Ref : Toxicon , 34 :959 , 1996
Abstract : Acetylcholinesterases (AChEs) very sensitive to fasciculin inhibition (KiS in picomolar range) have a distinctive group of aromatic amino acids in the peripheral region (Y70, Y121, W279 in Torpedo AChE). Enzymes that lack these amino acids like butyrylcholinesterases (BChEs) or one or two of them like cobra venom, insect and chicken AChEs are 1000 to 1,000,000 times less sensitive. Fasciculin is a non-competitive inhibitor of the hydrolysis of choline and neutral esters by very sensitive AChEs. For the other group of enzymes, differences arise according to the type of substrate. Fasciculin still behaves as a non-competitive inhibitor with choline esters. In contrast, hydrolysis of phenylacetate was unaffected or slightly increased with BChEs and a partial competitive inhibition was observed with cobra venom and chicken enzymes.
ESTHER : Duran_1996_Toxicon_34_959
PubMedSearch : Duran_1996_Toxicon_34_959
PubMedID: 8875783

Title : Purification and sequence determination of a new muscarinic toxin (MT4) from the venom of the green mamba (Dendroaspis angusticeps) - Vandermeers_1995_Toxicon_33_1171
Author(s) : Vandermeers A , Vandermeers-Piret MC , Rathe J , Waelbroeck M , Jolkkonen M , Oras A , Karlsson E
Ref : Toxicon , 33 :1171 , 1995
Abstract : A toxin which partially inhibited [3H]N-methylscopolamine binding to rat brain muscarinic receptors was purified from the venom of green mamba, Dendroaspis angusticeps. The N-terminal sequence (up to 45 amino acids) was determined by automated Edman degradation of the whole molecule. The complete sequence was elucidated after enzymatic cleavage with endoproteinase Arg-C or endoproteinase Lys-C and peptide fragments purification. The identity of the C-terminal amino acid was confirmed by hydrazinolysis. The new toxin (MT4) had eight half-cystines and 66 amino acids. It differed from muscarinic toxin MT1 by a single substitution in position 57 (arginine in MT1, histidine in MT4), proximal to the sixth half-cystine.
ESTHER : Vandermeers_1995_Toxicon_33_1171
PubMedSearch : Vandermeers_1995_Toxicon_33_1171
PubMedID: 8585087

Title : Role of arginine residues for the activity of fasciculin - Cervenansky_1995_Eur.J.Biochem_229_270
Author(s) : Cervenansky C , Engstrom A , Karlsson E
Ref : European Journal of Biochemistry , 229 :270 , 1995
Abstract : The West African green mamba, Dendroaspis angusticeps, has two toxins, fasciculins, that are non-competitive inhibitors of acetylcholinesterase. Arginine residues of fasciculin 2 were modified with 1,2-cyclohexanedione. Two of these residues, Arg24 and Arg37, reacted very slowly or not at all. Modification of Arg28 reduced the activity only by 13%. Arg11 and Arg27 are unique for fasciculins; a comparison of the sequences of 175 snake toxins homologous to fasciculins showed that no other toxin has arginine in the corresponding positions. Modification of the two unique arginines had a large effect and decreased the activity by 73% (Arg11) and 85% (Arg27). This was apparently not due to structural perturbations, since the modification did not change the circular dichroic spectra. The two arginine residues probably participate in the binding to acetylcholinesterase. They are located on the same side of the toxin molecule and the distance between their alpha-carbons is 2.7 nm. This may indicate binding to sites that are far apart and suggests that fasciculin covers a large area of the enzyme.
ESTHER : Cervenansky_1995_Eur.J.Biochem_229_270
PubMedSearch : Cervenansky_1995_Eur.J.Biochem_229_270
PubMedID: 7744040

Title : Poster: Distribution of m4 muscarinic cholinergic receptors in the rat brain detected by a selective mamba toxin (MT-3) -
Author(s) : Adem A , Bogdanovic N , Islam A , Jolkkonen M , Winblad B , Karlsson E
Ref : Life Sciences , 56(11-12) :1013 , 1995
PubMedID:

Title : A snake toxin against muscarinic acetylcholine receptors: amino acid sequence, subtype specificity and effect on guinea-pig ileum - Jolkkonen_1995_Toxicon_33_399
Author(s) : Jolkkonen M , Adem A , Hellman U , Wernstedt C , Karlsson E
Ref : Toxicon , 33 :399 , 1995
Abstract : The sequence of muscarinic toxin 1 (MT1) from Dendroaspis angusticeps (green mamba) was determined (66 amino acids, M(r) 7509). The central part, peptide 25-40, is rich in hydrophobic amino acids, which is a characteristic of muscarinic toxins. MT1 started to inhibit [3H]-NMS (N-methylscopolamine) binding to synaptosomal membranes of porcine brain (contains all five receptor subtypes) at about 1 nM and to membranes from pig heart muscle (only subtype m2) at about 1 microM. Binding of [3H]-AF-DX 384 to heart was inhibited with an IC50 of 14 microM and to brain in two steps. In the first step (IC50 = 32 nM) binding decreased by 37%, indicating that the toxin acted on m1 or m4 receptors, each accounting for about 40% of total receptor content. The second step was similar to the effect on heart. Pirenzepine inhibited binding of [125I]-MT1 to brain receptors with an IC50 of 6.5 nM, corresponding to a Ki of about 6 nM. Literature values of Ki for pirenzepine are 16-18 nM for m1 and > or = 120 mM for other subtypes. This indicates binding to m1 receptors. mM for other subtypes. This indicates binding to m1 receptors. [125I]-MT1 bound to brain with a Kd of 20 nM and a Hill coefficient of 1.0, i.e. one toxin molecule per receptor. In guinea-pig ileum, MT1 (670 nM) produced a rapid contraction, reversible by atropine. The toxin may be an agonist, but might also cause contraction by inducing acetylcholine release by a different mechanism.
ESTHER : Jolkkonen_1995_Toxicon_33_399
PubMedSearch : Jolkkonen_1995_Toxicon_33_399
PubMedID: 7570626

Title : Muscarinic toxins from the black mamba Dendroaspis polylepis - Jolkkonen_1995_Eur.J.Biochem_234_579
Author(s) : Jolkkonen M , van Giersbergen PL , Hellman U , Wernstedt C , Oras A , Satyapan N , Adem A , Karlsson E
Ref : European Journal of Biochemistry , 234 :579 , 1995
Abstract : Three new toxins acting on muscarinic receptors were isolated from the venom of the black mamba Dendroaspis polylepis. They were called muscarinic toxins alpha, beta, and gamma (MT alpha, MT beta, and MT gamma). All of the toxins have four disulphide bonds and 65 or 66 amino acids. The sequences of MT alpha and MT beta were determined. The muscarinic toxins, of which about 12 have been isolated from venoms of green and black mambas, have 60-98% sequence identity with each other, and are similar to many (about 180) other snake venom components, such as alpha-neurotoxins, cardiotoxins, and fasciculins. In contrast to the alpha-neurotoxins, muscarinic toxins do not bind to nicotinic acetylcholine receptors. The binding constants of MT alpha and MT beta were determined for human muscarinic receptors of subtypes m1-m5 stably expressed in Chinese hamster ovary cells. The toxins are less selective than the earlier discovered muscarinic toxins from the green mamba Dendroaspis angusticeps. MT alpha and the muscarinic toxin MT4 from D. angusticeps differ only in a region of three amino acids (residues 31-33), which are Leu-Asn-His in MT alpha and Ile-Val-Pro in MT4. This difference causes a pronounced shift in subtype selectivity. MT alpha has high affinity to all subtypes, with Ki (inhibition constant) values of 23 nM (m1; pKi = 7.64 +/- 0.10), 44 nM (m2; pKi = 7.36 +/- 0.06), 3 nM (m3; pKi = 8.46 +/- 0.14), 5 nM (m4; pKi = 8.32 +/- 0.07), and 8 nM (m5; pKi = 8.09 +/- 0.07). MT4 has high affinity only to m1 (Ki = 62 nM) and m4 (87 nM) receptors, and low (Ki > 1 microM) affinity to m2, m3, and m5. The region at positions 31-33 evidently plays an important role in the toxin-receptor interaction. MT beta has low affinity for m1 and m2 receptors (Ki > 1 microM) and intermediate affinity for m3 (140 nM; pKi = 6.85 +/- 0.03), m4 (120 nM; pKi = 6.90 +/- 0.06), and m5 (350 nM; pKi = 6.46 +/- 0.01). The low affinity of MT beta may reflect a tendency for spontaneous inactivation.
ESTHER : Jolkkonen_1995_Eur.J.Biochem_234_579
PubMedSearch : Jolkkonen_1995_Eur.J.Biochem_234_579
PubMedID: 8536706

Title : Protein toxins that bind to muscarinic acetylcholine receptors -
Author(s) : Karlsson E , Jolkkonen M , Satyapan N , Adem A , Kumlin E , Hellman U , Wernstedt C
Ref : Annals of the New York Academy of Sciences , 710 :153 , 1994
PubMedID: 8154745

Title : Two-step binding of green mamba toxin to muscarinic acetylcholine receptor - Toomela_1994_FEBS.Lett_352_95
Author(s) : Toomela T , Jolkkonen M , Rinken A , Jarv J , Karlsson E
Ref : FEBS Letters , 352 :95 , 1994
Abstract : The mechanism of binding of toxin MT2 from venom of green mamba Dendroaspis angusticeps to muscarinic acetylcholine receptors from rat cerebral cortex was investigated by studying the kinetics of the toxin-receptor interaction. The muscarinic antagonist N-methyl-[3H]scopolamine was used as a 'reporter' ligand. Evidence for a mechanism of toxin-receptor interaction comprising at least two steps was obtained. Such a mechanism increases the potency of the toxin. The first step was fast with no competition between the toxin and the antagonist. The second step was slow with formation of a more stable toxin-receptor complex and inhibition of the antagonist binding. It is proposed that the snake toxin is a muscarinic agonist of slow action.
ESTHER : Toomela_1994_FEBS.Lett_352_95
PubMedSearch : Toomela_1994_FEBS.Lett_352_95
PubMedID: 7925953

Title : Study of structure-activity relationship of fasciculin by acetylation of amino groups - Cervenansky_1994_Biochim.Biophys.Acta_1199_1
Author(s) : Cervenansky C , Engstrom A , Karlsson E
Ref : Biochimica & Biophysica Acta , 1199 :1 , 1994
Abstract : Dendroaspis angusticeps (green mamba) has two toxins, fasciculins, that are non-competitive inhibitors of acetylcholinesterase. Amino groups of fasciculin 2 were acetylated with acetic anhydride. The monoacetyl derivatives of the epsilon-amino groups (Lys 25, 32, 51 and 58) retained between 28 and 43% of the initial activity and that of the alpha-amino group 72%. Acetylation of Lys 25 that has the most reactive amino group decreased the activity by 65% apparently without producing structural perturbations, since the circular dichroism spectrum was not affected. The three-dimensional structure shows a cationic cluster formed by Lys 32, 51, Arg 24 and 28. A comparison of 175 sequences of homologous toxins shows that Lys 32 is unique for fasciculin. Acetylation of lysine residues in the cluster had a large effect and reduced the activity by 72% (Lys 32) and 57% (Lys 51). This suggests an important role for the cationic cluster. Lys 25 together with Lys 32 and 51 were, therefore, assumed to be in the active site.
ESTHER : Cervenansky_1994_Biochim.Biophys.Acta_1199_1
PubMedSearch : Cervenansky_1994_Biochim.Biophys.Acta_1199_1
PubMedID: 8280746

Title : A toxin from the green mamba Dendroaspis angusticeps: amino acid sequence and selectivity for muscarinic m4 receptors - Jolkkonen_1994_FEBS.Lett_352_91
Author(s) : Jolkkonen M , van Giersbergen PL , Hellman U , Wernstedt C , Karlsson E
Ref : FEBS Letters , 352 :91 , 1994
Abstract : Muscarinic toxin 3 (MT3) (65 amino acids, four disulphides, M(r) 7379) was isolated from the venom of the African snake Dendroaspis angusticeps (green mamba) and its amino acid sequence determined. Its ability to inhibit the binding of [3H]N-methylscopolamine ([3H]NMS) to Chinese hamster ovary cells stably expressing subtypes of muscarinic receptors was studied. MT3 displayed high affinity for the m4 receptor (pKi = 8.7 +/- 0.06), 40-fold lower affinity at ml receptors (pKi = 7.11 +/- 0.17) whereas no inhibition of [3H]NMS binding to m2, m3 and m5 receptors was observed at concentrations up to 1 microM. This makes MT3 the most selective m4 receptor ligand known to date.
ESTHER : Jolkkonen_1994_FEBS.Lett_352_91
PubMedSearch : Jolkkonen_1994_FEBS.Lett_352_91
PubMedID: 7925952

Title : Poster: Muscarinic toxin 1 (MT-1) receptors in the rat brain: Quantitative autoradiographic localization -
Author(s) : Adem A , Bogdanovic N , Karlsson E
Ref : Life Sciences , 52(5-6) :584 , 1993
PubMedID:

Title : Amino acid sequence of a snake venom toxin that binds to the muscarinic acetylcholine receptor - Karlsson_1991_Toxicon_29_521
Author(s) : Karlsson E , Risinger C , Jolkkonen M , Wernstedt C , Adem A
Ref : Toxicon , 29 :521 , 1991
Abstract : The green mamba, Dendroaspis angusticeps, has two protein toxins that bind to the muscarinic acetylcholine receptor. The sequence of muscarinic toxin 2 was determined with an automatic gas phase sequencer. The C-terminal residue is Asp as determined by hydrazinolysis and amino acid analysis. Toxin 2 has 65 amino acid residues and a formula weight of 7040. It is homologous to a large number of other snake venom toxins as short alpha-neurotoxins, cardiotoxins/cytotoxins and angusticeps-type toxins of mamba venoms. The sequence is confirmed in the accompanying article (Ducancel, F., Rowan, E.G., Cassart, E., Harvey, A. L., Menez, A. and Boulain, J.-C. Toxicon 29, 516-520, 1991).
ESTHER : Karlsson_1991_Toxicon_29_521
PubMedSearch : Karlsson_1991_Toxicon_29_521
PubMedID: 1862525

Title : Toxins from the venom of the green mamba Dendroaspis angusticeps that inhibit the binding of quinuclidinyl benzilate to muscarinic acetylcholine receptors - Adem_1988_Biochim.Biophys.Acta_968_340
Author(s) : Adem A , Asblom A , Johansson G , Mbugua PM , Karlsson E
Ref : Biochimica & Biophysica Acta , 968 :340 , 1988
Abstract : Two protein toxins that displace the muscarinic antagonist quinuclidinyl benzilate from rat cortex synaptosomal membranes have been isolated from the green mamba (Dendroaspis angusticeps) venom by gel filtration on sephadex G-50, chromatography on the ion-exchangers Bio-Rex 70 and Sulphopropyl-Sephadex C-25 and reversed-phase HPLC. Toxin 1 has 64 amino acids and four disulfides and a formula weight of 7200 and the corresponding values for toxin 2 are 63, 4 and 6840, respectively. Ultracentrifugation gave a molecular weight of 6900 for toxin 1 and 6700 for toxin 2, Quinuclidinyl benzilate that binds to all types of muscarinic cholinergic receptor was displaced to about 50% by both toxins. This partial displacement indicates that the toxins might be specific for one subtype of receptor.
ESTHER : Adem_1988_Biochim.Biophys.Acta_968_340
PubMedSearch : Adem_1988_Biochim.Biophys.Acta_968_340
PubMedID: 3345316

Title : Anticholinesterase toxins -
Author(s) : Karlsson E , Mbugua PM , Rodriguez-Ithurralde D
Ref : Pharmacol Ther , 30 :259 , 1985
PubMedID: 3842891

Title : Fasciculins, anticholinesterase toxins from the venom of the green mamba Dendroaspis angusticeps - Karlsson_1984_J.Physiol.(Paris)_79_232
Author(s) : Karlsson E , Mbugua PM , Rodriguez-Ithurralde D
Ref : Journal de Physiologie (Paris) , 79 :232 , 1984
Abstract : Two toxins that are potent inhibitors of acetylcholinesterase have been isolated from the venom of the green mamba, Dendroaspis angusticeps. The toxins have been called fasciculins since after injection into mice (i.p. 0.5-3 micrograms/g body weight) they cause severe, generalized and long-lasting (5-7 h) fasciculations. Homogenates of diaphragm, tibialis anterior and gastrocnemius muscles from mice injected with fasciculins showed a decrease in acetylcholinesterase activity by 45-60% compared to muscles from control animals. Histochemical staining revealed a greatly reduced acetylcholinesterase activity at neuromuscular junctions. Fasciculins have 61 amino acid residues and four disulfides. The molecular weights are 6765 (fasciculin 1) and 6735 (fasciculin 2). The sequences of the two toxins differ probably only at one position by a replacement of Tyr with Asp/Asn. 1 g of venom contained about 40 mg of fasciculins, 2/3 of which was fasciculin 2. A similar inhibitor has also been isolated from D. polylepis (black mamba) venom. The sequence of fasciculin 2 is known. Most of the positive charges are concentrated in a small section of the central part of the molecule, and most of the negative charges are in the C-terminal region. Fasciculins appear to have a pronounced dipole character. Fasciculin binds to the peripheral anionic site, since it can displace propidium, a probe for that site, from acetylcholinesterase. In vitro, in Krebs-Henseleit solution containing 2 mM NaH2PO4 (pH 7.4), fasciculin 2 inhibits acetylcholinesterase from human erythrocytes (Ki = 1.1 X 10(-10) M, 37 degrees C), rat muscle (Ki = 1.2 X 10(-10) M, 37 degrees C) and Electrophorus electricus (Ki = 3.0 X 10(-10) M, 22 degrees C).(ABSTRACT TRUNCATED AT 250 WORDS)
ESTHER : Karlsson_1984_J.Physiol.(Paris)_79_232
PubMedSearch : Karlsson_1984_J.Physiol.(Paris)_79_232
PubMedID: 6530667

Title : Protease inhibitor homologues from mamba venoms: facilitation of acetylcholine release and interactions with prejunctional blocking toxins - Harvey_1982_Br.J.Pharmacol_77_153
Author(s) : Harvey AL , Karlsson E
Ref : British Journal of Pharmacology , 77 :153 , 1982
Abstract : 1 Five polypeptides, which were isolated from elapid snake venoms and which are structurally related to protease inhibitors, were tested for action on isolated biventer cervicis nerve-muscle preparations of the chick. 2 Dendrotoxin from the Eastern green mamba (Dendroaspis angusticeps) and toxins K and I from the black mamba (Dendroaspis polylepis polylepis) increased to indirect stimulation without affecting responses to exogenous acetylcholine, carbachol of KCl. 3 The two other protease inhibitor homologues, HHV-II from Ringhals cobra (Hemachatus haemachatus) and NNV-II from Cape cobra (Naja nivea) did not increase responses to nerve stimulation. Trypsin inhibitor from bovine pancreas also had no facilitatory effects on neuromuscular transmission. 4 The facilitatory toxins from mamba venoms interacted with the prejunctional blocking toxins, beta-bungarotoxin, crotoxin and notexin, but not with taipoxin. The blocking effects of beta-bungarotoxin were reduced by pretreatment with the mamba toxins, whereas the blocking actions of crotoxin and notexin were enhanced. 5 The results indicate that protease inhibitor homologues from mamba venoms form a new class of neurotoxin, which acts to increase the release of acetylcholine in response to motor nerve stimulation. 6 From the interaction studies it is concluded that the facilitatory toxins bind to motor nerve terminals at sites related to those occupied by the prejunctional blocking toxins. However, differences in interactions with individual toxins suggest that there must be several related binding sites on the nerve terminals.
ESTHER : Harvey_1982_Br.J.Pharmacol_77_153
PubMedSearch : Harvey_1982_Br.J.Pharmacol_77_153
PubMedID: 6751453

Title : Interaction surfaces of neurotoxins and acetylcholine receptor - Tsetlin_1982_Toxicon_20_83
Author(s) : Tsetlin VI , Karlsson E , Utkin YN , Pluzhnikov KA , Arseniev AS , Surin AM , Kondakov VV , Bystrov VF , Ivanov VT , Ovchinnikov Yu A
Ref : Toxicon , 20 :83 , 1982
Abstract : Binding of neurotoxin II Naja naja oxiana derivatives containing one spin label at various positions (Leu 1, Glu 2, Lys 15, Lys 25, Lys 26, His 31, Lys 44 and Lys 46) to purified solubilized acetylcholine receptor protein (AchR) from Torpedo marmorata was studied by EPR techniques. AchR interaction with several dansylated neurotoxin II derivatives was followed by difference fluorescence spectroscopy. A series of neurotoxin II p-azidobenzoyl derivatives were prepared and in three of them modified lysine residues were identified. In combination, spectroscopic data and photolabeling implicate a considerable area of the neurotoxin in association with AchR. Rigidity of the neurotoxin II conformation allowed to regard its binding surface as a mould of the AchR corresponding site and to estimate the minimal size of the latter. Conformation of the long-chain neurotoxins and their binding to AchR are briefly discussed basing on the 1H and 19F NMR studies of neurotoxin I Naja naja oxiana, toxin 3 Naja naja siamensis and its acetylated or trifluoroacetylated derivatives, as well as on Achr interaction with the derivatives spin labeled at Lys 27 and His 71.
ESTHER : Tsetlin_1982_Toxicon_20_83
PubMedSearch : Tsetlin_1982_Toxicon_20_83
PubMedID: 7080049

Title : Dendrotoxin from the venom of the green mamba, Dendroaspis angusticeps. A neurotoxin that enhances acetylcholine release at neuromuscular junction -
Author(s) : Harvey AL , Karlsson E
Ref : Naunyn Schmiedebergs Arch Pharmacol , 312 :1 , 1980
PubMedID: 7393344

Title : EPR and fluorescence study of interaction of Naja naja oxiana neurotoxin II and its derivatives with acetylcholine receptor protein from Torpedo marmorata -
Author(s) : Tsetlin VI , Karlsson E , Arseniev AS , Utkin YN , Surin AM , Pashkov VS , Pluzhnikov KA , Ivanov VT , Bystrov VF , Ovchinnikov YA
Ref : FEBS Letters , 106 :47 , 1979
PubMedID: 227728

Title : Further studies on the binding properties of cobra neurotoxin to cholinergic receptors in mouse skeletal muscle - Libelius_1975_J.Neural.Transm_37_165
Author(s) : Libelius R , Eaker D , Karlsson E
Ref : J Neural Transm , 37 :165 , 1975
Abstract : Preparations of 3H-monoacetylated derivatives of Naja naja siamensis neurotoxin siamensis 3 were found to be resistant to degradation and deacetylation during in vitro muscle incubation. A low rate of free 3H-acetate (less than 0.4%) was present in the preparations, but should not interfere with the binding of toxin to cholinergic receptors in mouse extensor digitorum longus muscles in vitro. The binding of toxin to cholinergic receptors in mouse extensor digitorum longus muscle was essentially irreversible. d-Tubocurarine antagonized the binding of toxin in both innervated and denervated muscles. Administration of actinomycin D one day after denervation prevented the appearance of new toxin binding sites in the muscles.
ESTHER : Libelius_1975_J.Neural.Transm_37_165
PubMedSearch : Libelius_1975_J.Neural.Transm_37_165
PubMedID: 1185161

Title : [Biological activities of various fractions isolated from venom of Hemachatus haemachates] -
Author(s) : Cheymol J , Karlsson E , Bourillet F , Roch-Arveiller M
Ref : Archives Internationales de Pharmacodynamie et de Therapie , 208 :81 , 1974
PubMedID: 4846476

Title : Effects of an isolated toxin from Australian tiger snake (Notechis scutatus scutatus) venom at the mammalian neuromuscular junction - Harris_1973_Br.J.Pharmacol_47_141
Author(s) : Harris JB , Karlsson E , Thesleff S
Ref : British Journal of Pharmacology , 47 :141 , 1973
Abstract : 1. The acute effects of a purified toxin from Australian Tiger snake (Notechis scutatus scutatus) venom have been investigated at the mammalian neuromuscular junction.2. The toxin was injected into the tail vein of mice. Death was due to respiratory paralysis.3. The resting membrane potential, and action potential of muscle fibres in muscles from in vivo intoxicated animals were normal.4. The frequency of miniature end plate potentials (m.e.p.p.s) from intoxicated nerve-muscle preparations was reduced, although m.e.p.p. amplitude was unaltered.5. Nerve stimulation resulted in end plate potentials (e.p.p.s) of quantal amplitude; only rarely was the e.p.p. large enough to give rise to an action potential.6. High (20 mM) K(+) did not increase m.e.p.p. frequency in intoxicated preparations.7. The toxin was largely ineffective in vitro.8. The similarities and differences between this toxin, beta-bungarotoxin and botulinum toxin are discussed.
ESTHER : Harris_1973_Br.J.Pharmacol_47_141
PubMedSearch : Harris_1973_Br.J.Pharmacol_47_141
PubMedID: 4352085

Title : Isolation of the nicotinic acetylcholine receptor by biospecific chromatography on insolubilized Naja naja neurotoxin -
Author(s) : Karlsson E , Heilbronn E , Widlund L
Ref : FEBS Letters , 28 :107 , 1972
PubMedID: 4646866

Title : Purification of a presynaptic neurotoxin from the venom of the australian tiger snake Notechis scutatus scutatus -
Author(s) : Karlsson E , Eaker D , Ryden L
Ref : Toxicon , 10 :405 , 1972
PubMedID: 5070579