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Unformatted text preview: ular loop that connects the III and IV transmembrane segments, as shown in Fig. 6. Presumably pronase acted by cutting some site or sites on that intracellular loop. 6 Figure 6. Two views of the operation of the inactivation gate. The top panel shows the ball and chain view, where the ball (shown as an oval) is attached by two linkers to two segments of the channel. The idea is that when the voltage sensor in S4 moves in response to depolarization and opens the activation gate (open state in middle), ions flow through the pore. A moment later the ball is pulled into the pore, blocking it and preventing any further flow of ions, even though the cell is still depolarized and the activation gate (pore) is "open". The bottom panel shows this in more detail. Specifically, the linkers connect to only two domains, domain III and domain IV (domain II is not shown for clarity). The ball is formed by the IFM motif of amino acids, with one side linked to domain III and the other to domain IV (IFM is a 3 amino acid sequence, isoleucine (I), phenylalanine (F), and methionine (M)). When the cell is depolarized, the linker in domain IV is pulled upward and causes the ball, formed by the IFM sequence, to plug the channel thereby producing channel inactivation. Thee four domains are shown, but the segments in each domain are not shown. The linkage from domain III is on S6 while the linkage on domain 4 is on S1. The linkages to each segment in domains III and IV can be seen in Fig. 2 and 7. There is only one inactivation gate on sodium channels. The gate consists of a ball and two chains. The ball has a unique amino acid sequence, the IFM motif, that is connected to intracellular linkages, the chains (Fig. 6). One chain connects the ball to the S6 segment in domain III while the other connects the ball to the S1 segment in domain IV (see Figs. 6, 7). Here is how it works. Upon depolarization, the voltage sensors (S4 segments) in domains I, II and III move and open the activation gate. Na+ ions now flows through the channel pore, from the outside of the cell to the inside, thereby generating the upstroke of the action potential. The voltage sensor (S4 segment) in domain IV, however, is slower than the voltage sensors in the other domains, and begins to move about a millisecond after the other voltage sensors have already moved. When the voltage sensor in domain IV is displaced upward, it moves the ball and causes it to block the channel pore. Thus, although the cell remains depolarized and the activation gates remain open, current now stops because the pore is blocked by the inactivation ball. The channel is now inactivated. Thus, in the Na+ channel the four domains do not behave equally. The inactivation complex interacts with the segments of domains III and IV but is not affected by domains I and II. The activation gate is controlled entirely by the S4 segments in domains I, II and III, while the inactivation gate is controlled by the S4 segment in domain 4. 7 The pore region The pore, the part of the channel through which ions flow from the extracellular to the intracellular fluids, is formed by S56. The amino acids linking the two segments actually dip into the pore and form the selectivity filter, the structure that enables the channel to selectively pass Na+ ions and excludes all other ions. The detailed arrangement of the pore and the way the selectivity filter operates has been worked out for K+ channels and is considered in a later section. Figure 7. Differential movements of S4 segments in the four domains of the Na+ channel produce activation and inactivation. The structure of potassium channels The first potassium channel to be sequenced came from the fruit fly, Drosophila melanogaster. Drosophila has been an invaluable resource for genetic studies and has made critically important contributions to our understanding of how K+ channels operate. Of special importance is the Shaker mutant. Researchers use ether as an anesthetic to count various mutants during their screens (otherwise the it just flies away). Occasionally, instead of just going to sleep, some flies shake the...
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- Spring '08