Mechanism and molecular basis for the sodium channel subtype specificity of μ-conopeptide CnIIIC
Mechanism and molecular basis for the sodium channel subtype specificity of μ-conopeptide CnIIIC
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DOI：
10.1111/j.1476-5381.2012.02004.x
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发表时间：
2012-10-01
影响因子：
7.3
通讯作者：
Heinemann, Stefan H.
中科院分区：
文献类型：
--
作者：
Markgraf, Rene;Leipold, Enrico;Heinemann, Stefan H.
背景和目的电压门控钠通道（NaV通道）在动作电位的产生和传播中起关键作用，选择性阻断这些通道是临床上有用的电活动抑制的有前景的策略。来自锥形蜗牛Conus consors的芋螺毒素mu-CnIIIC通过特异性阻断骨骼肌（NaV 1.4）NaV通道在啮齿动物中表现出肌松弛活性。实验方法我们研究了mu-CnIIIC对人NaV通道的活性，并表征了其抑制机制以及其通道特异性的分子基础。关键结果与大鼠旁系同源物相似，人NaV1.4和NaV1.2被mu-CnIIIC有效阻断，NaV1.7的敏感性居中，NaV1.5和NaV1.8不敏感。半通道嵌合体显示，决定因素的不敏感性的NaV1.8必须驻留在第一和第二半的通道，而那些NaV1.5仅限于域I和II。此外，结构域I孔环影响总块，因此窝藏亚型特异性的主要决定因素。结构域II孔环仅影响毒素结合和解离的动力学。Mu-CnIIIC对NaV1.4的阻断几乎是不可逆的，但留下了约5%的残余电流，反映了泄漏阻断;因此，Na+离子在一定程度上仍然通过Mu-CnIIIC占据的NaV1.4。TTX被排除在该结合位点之外，但被mu-CnIIIC捕获在孔内。结论和意义的临床意义，mu-CnIIIC是一种有效的和持久的人骨骼肌NaV1.4的阻滞剂，不影响心脏NaV1.5的活动。
BACKGROUND AND PURPOSE Voltage-gated sodium channels (NaV channels) are key players in the generation and propagation of action potentials, and selective blockade of these channels is a promising strategy for clinically useful suppression of electrical activity. The conotoxin mu-CnIIIC from the cone snail Conus consors exhibits myorelaxing activity in rodents through specific blockade of skeletal muscle (NaV1.4) NaV channels. EXPERIMENTAL APPROACH We investigated the activity of mu-CnIIIC on human NaV channels and characterized its inhibitory mechanism, as well as the molecular basis, for its channel specificity. KEY RESULTS Similar to rat paralogs, human NaV1.4 and NaV1.2 were potently blocked by mu-CnIIIC, the sensitivity of NaV1.7 was intermediate, and NaV1.5 and NaV1.8 were insensitive. Half-channel chimeras revealed that determinants for the insensitivity of NaV1.8 must reside in both the first and second halves of the channel, while those for NaV1.5 are restricted to domains I and II. Furthermore, domain I pore loop affected the total block and therefore harbours the major determinants for the subtype specificity. Domain II pore loop only affected the kinetics of toxin binding and dissociation. Blockade by mu-CnIIIC of NaV1.4 was virtually irreversible but left a residual current of about 5%, reflecting a leaky block; therefore, Na+ ions still passed through mu-CnIIIC-occupied NaV1.4 to some extent. TTX was excluded from this binding site but was trapped inside the pore by mu-CnIIIC. CONCLUSION AND IMPLICATIONS Of clinical significance, mu-CnIIIC is a potent and persistent blocker of human skeletal muscle NaV1.4 that does not affect activity of cardiac NaV1.5.