Ji S, George A L, Horn R, Barchi R L
Departments of Neurology and Neuroscience, University of Pennsylvania School of Medicine, Philadelphia, 19104-6074, USA.
J Gen Physiol. 1996 Feb;107(2):183-94. doi: 10.1085/jgp.107.2.183.
Mutations in the gene encoding the voltage-gated sodium channel of skeletal muscle (SkMl) have been identified in a group of autosomal dominant diseases, characterized by abnormalities of the sarcolemmal excitability, that include paramyotonia congenita (PC) and hyperkalemic periodic paralysis (HYPP). We previously reported that PC mutations cause in common a slowing of inactivation in the human SkMl sodium channel. In this investigation, we examined the molecular mechanisms responsible for the effects of L1433R, located in D4/S3, on channel gating by creating a series of additional mutations at the 1433 site. Unlike the R1448C mutation, found in D4/S4, which produces its effects largely due to the loss of the positive charge, change of the hydropathy of the side chain rather than charge is the primary factor mediating the effects of L1433R. These two mutations also differ in their effects on recovery from inactivation, conditioned inactivation, and steady state inactivation of the hSkMl channels. We constructed a double mutation containing both L1433R and R1448C. The double mutation closely resembled R1448C with respect to alterations in the kinetics of inactivation during depolarization and voltage dependence, but was indistinguishable from L1433R in the kinetics of recovery from inactivation and steady state inactivation. No additive effects were seen, suggesting that these two segments interact during gating. In addition, we found that these mutations have different effects on the delay of recovery from inactivation and the kinetics of the tail currents, raising a question whether this delay is a reflection of the deactivation process. These results suggest that the S3 and S4 segments play distinct roles in different processes of hSkM1 channel gating: D4/S4 is critical for the deactivation and inactivation of the open channel while D4/S3 has a dominant role in the recovery of inactivated channels. However, these two segments interact during the entry to, and exit from, inactivation states.
在一组常染色体显性疾病中，已发现编码骨骼肌电压门控钠通道（SkMl）的基因突变，这些疾病的特征是肌膜兴奋性异常，包括先天性副肌强直（PC）和高钾性周期性麻痹（HYPP）。我们之前报道过，PC突变共同导致人类SkMl钠通道失活减慢。在本研究中，我们通过在1433位点创建一系列额外突变，研究了位于D4/S3的L1433R对通道门控作用的分子机制。与在D4/S4中发现的R1448C突变不同，后者主要由于正电荷丢失而产生效应，侧链亲水性的改变而非电荷是介导L1433R效应的主要因素。这两种突变对hSkMl通道失活恢复、条件性失活和稳态失活的影响也有所不同。我们构建了同时包含L1433R和R1448C的双突变体。该双突变体在去极化期间失活动力学和电压依赖性改变方面与R1448C非常相似，但在失活恢复动力学和稳态失活方面与L1433R无法区分。未观察到加性效应，表明这两个片段在门控过程中相互作用。此外，我们发现这些突变对失活恢复延迟和尾电流动力学有不同影响，这引发了一个问题，即这种延迟是否是失活过程的反映。这些结果表明，S3和S4片段在hSkM1通道门控的不同过程中发挥着不同作用：D4/S4对开放通道的失活和去激活至关重要，而D4/S3在失活通道的恢复中起主导作用。然而，这两个片段在进入和离开失活状态的过程中相互作用。
