IONIC MOVEMENTS ACROSS NERVE CELL BODY MEMBRANES

长期目标是指定电压传感器和孔 电压依赖性脑K+通道和失活门 电压依赖性脑Na+通道。 具体目标是：1） 建立用于所述单通道动力学的参考数据集，以及 电导的大鼠脑K+通道drk 1，我们最近有 克隆、测序和表达; 2）测试广泛变化的影响 在drk 1的非核心区中， 最小功能结构; 3）通过以下方式将结构与功能相关联： 比较密切相关的单通道电流 同形变体; 4）在区域中产生点突变或嵌合体 被认为是电压传感器或3C通道的传导孔或 Ne通道的失活门; 5）突变大鼠胞质接头 脑Na+II以测试其在失活门控中的作用。 在所有情况下的 三级河道结构使用基于已知的 一级氨基酸序列。 通过注射来检验假设。 将来自天然同种型和工程突变体的转录物转化为 非洲爪蟾卵母细胞。 从电压依赖性的比较， 在不同状态之间转换到闭合状态和打开状态以及从闭合状态和打开状态转换 蛋白质，可以推断门控结构。 通过比较 双离子溶液中的单通道电导， 结构可以推断。 结果及其解释将是 扩展到人脑K+和NC通道。 结构知识 负责电压门控和离子通量将提供合理的 治疗癫痫、帕金森综合症和 老年痴呆症。

The long-term objective is to specify the voltage sensor and the pore of voltage-dependent brain K+ channels and the inactivation gate of voltage-dependent brain Na+ channels. The specific aims are to: 1) establish a reference data set for the single channel kinetics and conductance of a rat brain K+ channel drk1, which we have recently cloned, sequenced and expressed; 2) test the effects of extensive changes in the non-core regions of drkl and engineer with recombinant methods a minimum functional structure; 3) correlate structure with function by comparing the single channels currents produced by closely related isomorphic variants; 4) produce point mutations or chimeras in regions thought to be the voltage sensor or conducting pore of 3C channels or the inactivation gate of Ne channels; 5) mutate the cytoplasmic linker in rat brain Na+II to test its role in inactivation gating. In all cases the tertiary channel structures are interpreted using models based upon known primary amino acid sequences. The hypotheses are tested by injecting transcripts from the natural isoforms and the engineered mutants into Xenopus oocytes. From a comparison of the voltage-dependence of transitions to and from the closed and open states among the different proteins, the gating structure may be inferred. From a comparison of single channel conductance in bi-ionic solutions, the permeation structure may be inferred. The results and their interpretation will be extended to human brain K+ and NC channels. Knowledge of the structures responsible for voltage gating and ionic flux will provide a rational basis for therapy of disorders such as epilepsy, Parkinsonism and Alzheimers disease.