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Chemical modification of phospholipase A2 from the venom of king brown snake, Pseudechis australis
First A s i a - Pacific Congress
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the toxin with the control bathing solution. Effects of EqT III on the nodal membrane conductances were less coherent. Destruction of the nodal membrane, seen as a huge and irreversible increase of the leakage conductance after application of EqT III in 1 - 100 nM concentration was the most regular event. Compared to the frog skeletal muscle fibres, there was no noticahle decrease of the sodium current amplitude after the treatment of nerve fibres with EqT, which might imply that either the structure of the sodium channel or the structure of the membrane is different in nerve and muscle fibres. Higher concentrations of EqT needed to produce an effect on nerves are probably due to the 'buffering' capacity of the myelin sheath for EqT. Results allow the conclusion that EqT I and EqT II are potent potassium channel blockers in the node.
Pathogenesis o f tetanus, clinically relevant new concepts. K. TAKANO and F. KIRCHNER (Department of Physiology and Pathophysiology, University of GOttingen, G6ttingen, F.R.G.). THIS STUDY gives a review of the pathogenesis of the hyperactivity of the motor system during tetanus intoxication primarily of our own experiments on animals. Tetanus toxin at doses of 1 - 100,000 mouse MLD/kg was injected into the muscle or vein of the cat, rabbit or rat for experimental local or general tetanus. Tetanus toxin blocked not only the inhibitory synapses of the spinal motoneurones as had been believed, but also the excitatory synapses at lower doses of tetanus toxin. At the minimal dose which caused local tetanus the monosynaptic reflex was always depressed or blocked. Presynaptic inhibition could be observed in local tetanus during the period when the monosynaptic reflex was present. Many types of spinal inhibition were left intact in general tetanus. The rhythmic activity of alpha as well as gamma motoneurones in the spinal cord in general tetanus could not be observed distally to the level of spinal transection. We conclude that clinically relevant tetanus is most likely of central rather than of spinal origin.
Chemical modification o f phospholipase A2 from the venom o f king brown snake, Pseudechis australis. C. TAKASAKI, A. YANAGITA, A. SUGAMAand N. TAMIYA (Department of Chemistry, Tohoku University, Sendal, Japan). Pseudechis australis phospholipase A2, Pa-I l, (phospholipase activity, 3030 units/mg protein; LDs00.23 gg/g of mouse) is chemically modified and its enzymic and lethal activities are studied. (1) Carbamoylation of lysyl residues. The enzyme protein (with 14 lysyl residues) is carbamoylated with K'~CNO and a mixture of monocarbamoyl derivatives is separated by chromatography on a CM-52 column. Mono-carbamoyl derivatives on 58-, 63-, 81- and 85- lysine residues are isolated and other derivatives obtained in mixtures. 63-carbamoyl lysine Pa-11 shows one tenth of the enzymic and lethal activities as compared to the original protein. 58-carbamoyl lysine Pa-11 has 30°70 higher enzymic activity and slightly lower lethal activity. 81- and 85-carbamoyl derivatives have the same activity as the original protein. (2) Carhoxymethylation of methionine. Pa-ll is carboxymethylated at its only methionine-8 with ICH2COOH in 6 M guanidine at pH 3.0. The derivative shows 7070 and less than 4070 of the enzymic and lethal activities, respectively. (3) Nitrophenylsulphenylation of tryptophan. Pa-I ! is treated with nitrophenylsulphenyl chloride. The modification resulted in complete loss of both enzymic and lethal activities. These results suggest that the enzymic activity is indispensable for the lethal activity, although there is no proportional relationship between the two activities.
Anticholinergic action o f a pedicellarial glycoprotein from the sea urchin Toxopneustes pileolus. M. TAKEI,~ K. ENDOH,l H. NAKAGAWA2 and A. KIMURA2 (~Department of Pharmacology, School of Pharmacy, Tokushima University of Arts and Science, and 2Department of Health Science, Faculty of Integrated Arts and Sciences, University of Tokushima, Tokushima 770, Japan). A PEDICELLARIALglycoprotein (con A fraction) from the sea urchin Toxopneustespileolus has been fractionated by a combination of gel chromatography, ion-exchange chromatography and lectin affinity chromatography. The purpose of this study was to characterize the pharmacological properties of con A fraction in the isolated skeletal muscle preparation. We have found that the pedicellarial extract and gel chromatographic fraction antagonize acetylcholine-inducedcontraction of frog rectus ahdominis. Con A fraction also had the same effect as these fractions. The inhibitory effect of con A fraction was approximately 10 times higher than that of gel chromatographic fraction. In the high K ÷-induced contraction, however, no appreciable effect of con A fraction was observed. Likwise, in the presence of AChE inhibitor, the effect of con A fraction was completely blocked. The present data suggest that the inhibitory action of con A fraction are mediated through the cholinergic receptor sites.
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the toxin with the control bathing solution. Effects of EqT III on the nodal membrane conductances were less coherent. Destruction of the nodal membrane, seen as a huge and irreversible increase of the leakage conductance after application of EqT III in 1 - 100 nM concentration was the most regular event. Compared to the frog skeletal muscle fibres, there was no noticahle decrease of the sodium current amplitude after the treatment of nerve fibres with EqT, which might imply that either the structure of the sodium channel or the structure of the membrane is different in nerve and muscle fibres. Higher concentrations of EqT needed to produce an effect on nerves are probably due to the 'buffering' capacity of the myelin sheath for EqT. Results allow the conclusion that EqT I and EqT II are potent potassium channel blockers in the node.
Pathogenesis o f tetanus, clinically relevant new concepts. K. TAKANO and F. KIRCHNER (Department of Physiology and Pathophysiology, University of GOttingen, G6ttingen, F.R.G.). THIS STUDY gives a review of the pathogenesis of the hyperactivity of the motor system during tetanus intoxication primarily of our own experiments on animals. Tetanus toxin at doses of 1 - 100,000 mouse MLD/kg was injected into the muscle or vein of the cat, rabbit or rat for experimental local or general tetanus. Tetanus toxin blocked not only the inhibitory synapses of the spinal motoneurones as had been believed, but also the excitatory synapses at lower doses of tetanus toxin. At the minimal dose which caused local tetanus the monosynaptic reflex was always depressed or blocked. Presynaptic inhibition could be observed in local tetanus during the period when the monosynaptic reflex was present. Many types of spinal inhibition were left intact in general tetanus. The rhythmic activity of alpha as well as gamma motoneurones in the spinal cord in general tetanus could not be observed distally to the level of spinal transection. We conclude that clinically relevant tetanus is most likely of central rather than of spinal origin.
Chemical modification o f phospholipase A2 from the venom o f king brown snake, Pseudechis australis. C. TAKASAKI, A. YANAGITA, A. SUGAMAand N. TAMIYA (Department of Chemistry, Tohoku University, Sendal, Japan). Pseudechis australis phospholipase A2, Pa-I l, (phospholipase activity, 3030 units/mg protein; LDs00.23 gg/g of mouse) is chemically modified and its enzymic and lethal activities are studied. (1) Carbamoylation of lysyl residues. The enzyme protein (with 14 lysyl residues) is carbamoylated with K'~CNO and a mixture of monocarbamoyl derivatives is separated by chromatography on a CM-52 column. Mono-carbamoyl derivatives on 58-, 63-, 81- and 85- lysine residues are isolated and other derivatives obtained in mixtures. 63-carbamoyl lysine Pa-11 shows one tenth of the enzymic and lethal activities as compared to the original protein. 58-carbamoyl lysine Pa-11 has 30°70 higher enzymic activity and slightly lower lethal activity. 81- and 85-carbamoyl derivatives have the same activity as the original protein. (2) Carhoxymethylation of methionine. Pa-ll is carboxymethylated at its only methionine-8 with ICH2COOH in 6 M guanidine at pH 3.0. The derivative shows 7070 and less than 4070 of the enzymic and lethal activities, respectively. (3) Nitrophenylsulphenylation of tryptophan. Pa-I ! is treated with nitrophenylsulphenyl chloride. The modification resulted in complete loss of both enzymic and lethal activities. These results suggest that the enzymic activity is indispensable for the lethal activity, although there is no proportional relationship between the two activities.
Anticholinergic action o f a pedicellarial glycoprotein from the sea urchin Toxopneustes pileolus. M. TAKEI,~ K. ENDOH,l H. NAKAGAWA2 and A. KIMURA2 (~Department of Pharmacology, School of Pharmacy, Tokushima University of Arts and Science, and 2Department of Health Science, Faculty of Integrated Arts and Sciences, University of Tokushima, Tokushima 770, Japan). A PEDICELLARIALglycoprotein (con A fraction) from the sea urchin Toxopneustespileolus has been fractionated by a combination of gel chromatography, ion-exchange chromatography and lectin affinity chromatography. The purpose of this study was to characterize the pharmacological properties of con A fraction in the isolated skeletal muscle preparation. We have found that the pedicellarial extract and gel chromatographic fraction antagonize acetylcholine-inducedcontraction of frog rectus ahdominis. Con A fraction also had the same effect as these fractions. The inhibitory effect of con A fraction was approximately 10 times higher than that of gel chromatographic fraction. In the high K ÷-induced contraction, however, no appreciable effect of con A fraction was observed. Likwise, in the presence of AChE inhibitor, the effect of con A fraction was completely blocked. The present data suggest that the inhibitory action of con A fraction are mediated through the cholinergic receptor sites.