Jerusalinsky D, Cerveñasky C, Peña C, Raskovsky S, Dajas F
Instituto de Biologia Celula, Facultad de Medicina, Universidad de Buenos Aires, Argentina.
Neurochem Int. 1992 Feb;20(2):237-46. doi: 10.1016/0197-0186(92)90173-o.
Two new polypeptides were isolated and purified from the venom of the snake Dendroaspis angusticeps, which also contains other neuroactive peptides such as Dendrotoxins and Fasciculins. The amino acid composition of the peptides was determined and the first 10 amino acids from the MTX2 N-terminal fragment were sequenced. The so-called muscarinic toxins (MTX1 and MTX2) have been shown to inhibit the specific binding of [3H]QNB (0.15 nM), [3H]PZ (2.5 nM) and [3H]oxoM (2 nM) to bovine cerebral cortex membranes by 60, 88 and 82% respectively. In contrast, they caused only a 30% blockade of the [3H]QNB specific binding to similar membrane preparations from the brainstem. The Hill number for the [3H]PZ binding inhibition by the putative muscarinic toxin MTX2 was 0.95 suggesting homogeneity in the behaviour of the sites involved. The data from [3H]oxoM binding gave a Hill number of 0.83. The decreases in the specific binding involved increases in KD for the three different ligands (8-fold for [3H]QNB, 4-fold for [3H]PZ and 3.5-fold for [3H]oxoM) without significant changes in Bmax, except for a slight decrease in the [3H]oxoM binding sites (-19%); such results suggest that there may be a competitive inhibition between the MTXs and these ligands. The Ki for MTX2/[3H]PZ was 22.58 +/- 3.52 nM; for MTX2/[3H]oxoM, 144.9 +/- 21.07 nM and for MTX2/[3H]QNB, 134.98 +/- 18.35 nM. The labelling of MTX2 with 125I allowed direct demonstration of specific and saturable binding to bovine cerebral cortex synaptosomal membranes. In conclusion, the results reported in this study strongly support the hypotheses that the two polypeptides isolated from D. angusticeps venom selectively inhibit specific ligand binding to central muscarinic receptors, in a competitive manner at least for the antagonist [3H]PZ and that the MTX2 specifically binds to a central site that is suggested to be a muscarinic receptor of the M1 subtype.
从黑曼巴蛇（Dendroaspis angusticeps）毒液中分离并纯化出了两种新的多肽，该毒液还含有其他神经活性肽，如树眼镜蛇毒素（Dendrotoxins）和束丝藻毒素（Fasciculins）。测定了这些肽的氨基酸组成，并对MTX2 N端片段的前10个氨基酸进行了测序。所谓的毒蕈碱毒素（MTX1和MTX2）已被证明可分别抑制[3H]QNB（0.15 nM）、[3H]PZ（2.5 nM）和[3H]氧代M（2 nM）与牛大脑皮层膜的特异性结合，抑制率分别为60%、88%和82%。相比之下，它们对[3H]QNB与脑干类似膜制剂的特异性结合仅造成30%的阻断。推测的毒蕈碱毒素MTX2对[3H]PZ结合抑制的希尔系数为0.95，表明所涉及位点的行为具有同质性。[3H]氧代M结合的数据给出的希尔系数为0.83。特异性结合的减少涉及三种不同配体的解离常数（KD）增加（[3H]QNB增加8倍，[3H]PZ增加4倍，[3H]氧代M增加3.5倍），最大结合量（Bmax）无显著变化，除了[3H]氧代M结合位点略有减少（-19%）；这些结果表明MTXs与这些配体之间可能存在竞争性抑制。MTX2/[3H]PZ的抑制常数（Ki）为22.58±3.52 nM；MTX2/[3H]氧代M为144.9±21.07 nM，MTX2/[3H]QNB为134.98±18.35 nM。用125I标记MTX2可直接证明其与牛大脑皮层突触体膜的特异性和饱和性结合。总之，本研究报告的结果有力地支持了以下假设：从黑曼巴蛇毒液中分离出的两种多肽选择性地抑制特异性配体与中枢毒蕈碱受体的结合，至少对于拮抗剂[3H]PZ是以竞争性方式，并且MTX2特异性结合到一个被认为是M1亚型毒蕈碱受体的中枢位点。
