Portal de Periódicos da CAPES

One Channel: Open and Closed

2005; Elsevier BV; Volume: 13; Issue: 10 Linguagem: Inglês

10.1016/j.str.2005.09.004

1878-4186

Janice Robertson, Benoı̂t Roux,

Cardiac electrophysiology and arrhythmias

Kuo et al., 2005Kuo, A., Domene, C., Johnson, L.N., Doyle, D.A., and Vénien-Bryan, C. (2005). Structure 13, this issue, 1463–1472.Google Scholar report structural information about the prokaryotic KirBac3.1 inward rectifier family K+ channel from Magnetospirillum magnetotacticum. These results from two-dimensional electron cryomicroscopy (EM) shed light on the gating mechanism of members of the Kir channel family. Kuo et al., 2005Kuo, A., Domene, C., Johnson, L.N., Doyle, D.A., and Vénien-Bryan, C. (2005). Structure 13, this issue, 1463–1472.Google Scholar report structural information about the prokaryotic KirBac3.1 inward rectifier family K+ channel from Magnetospirillum magnetotacticum. These results from two-dimensional electron cryomicroscopy (EM) shed light on the gating mechanism of members of the Kir channel family. K+ channels are large transmembrane proteins that control and regulate the passage of K+ ions across the cell membrane. Significant conformational changes of the protein are expected to underlie the process, referred to as "gating," by which a channel opens and closes its passageway to ions. To understand the specific gating mechanism for a given family of K+ channels, it is essential to determine the three-dimensional (3D) structure of at least two functional states of the same protein. In this issue of Structure, Kuo et al., 2005Kuo, A., Domene, C., Johnson, L.N., Doyle, D.A., and Vénien-Bryan, C. (2005). Structure 13, this issue, 1463–1472.Google Scholar report EM data for two conformational states of KirBac3.1 at 9 Å resolution, in which they observe a substantial increase in pore size from less than 6 Å to greater that 11 Å. The EM data provide a projection view of the KirBac3.1 channel in the natural environment of a lipid bilayer membrane. Depending on the experimental conditions, two different crystal forms were observed: one obtained in the presence of Mg2+, in which the channel structure appears tightly closed, and another in the absence of Mg2+, in which the channel structure is more expanded and putatively open. Using calculations of cross correlation values between the 2D EM projection data and the 3D atomic structure of Kirbac1.1 (a homolog of Kirbac3.1) that was previously determined by X-ray crystallography (Kuo et al., 2003Kuo A. Gulbis J.M. Antcliff J.F. Rahman T. Lowe E.D. Zimmer J. Cuthbertson J. Ashcroft F.M. Ezaki T. Doyle D.A. Science. 2003; 300: 1922-1926Crossref PubMed Scopus (704) Google Scholar), the authors were able to determine a structural model that fits well to the expanded state density. Although there is no direct evidence as to the functionality of this expanded conformation, all intuition suggests that this may be an open state of the channel. The analysis reveals two distinct conformational states for the same channel protein. To date, structural comparisons between open and closed states have been made only between channel families but not within: KcsA (Doyle et al., 1998Doyle D.A. Morais Cabral J. Pfuetzner R.A. Kuo A. Gulbis J.M. Cohen S.L. Chait B.T. MacKinnon R. Science. 1998; 280: 69-77Crossref PubMed Scopus (5466) Google Scholar) and KirBac1.1 (Kuo et al., 2003Kuo A. Gulbis J.M. Antcliff J.F. Rahman T. Lowe E.D. Zimmer J. Cuthbertson J. Ashcroft F.M. Ezaki T. Doyle D.A. Science. 2003; 300: 1922-1926Crossref PubMed Scopus (704) Google Scholar) for the closed state, and MthK (Jiang et al., 2002Jiang Y. Lee A. Chen J. Cadene M. Chait B.T. MacKinnon R. Nature. 2002; 417: 523-526Crossref PubMed Scopus (1043) Google Scholar) for the open state. While those comparisons reveal the overall nature of the conformational change underlying channel closing and opening—the inner helices form a constriction point that blocks the pore entrance in the closed state of the channel, whereas they are bent and splayed outward at a highly conserved glycine hinge residue in the open state—the structures provided by Kuo et al. allow the examination of this general K+ channel gating mechanism within the reference frame of a single channel type. In particular, these results put important constraints on the gating mechanism for members of the inward rectifier family of K+ channels (Kir). There has been considerable debate about the conformational changes associated with Kir channel gating. While one hypothesis proposes an intracellular gate similar to that of the helical bundle seen in KcsA, a number of other studies have suggested that changes in the selectivity filter itself may act as the predominant gate in these channels (Bichet et al., 2003Bichet D. Haass F.A. Jan L.Y. Nat. Rev. Neurosci. 2003; 4: 957-967Crossref PubMed Scopus (196) Google Scholar). Although the existence of additional gates remains possible, the previous closed state crystal structure of KirBac1.1, together with the new structural data by Kuo et al., 2005Kuo, A., Domene, C., Johnson, L.N., Doyle, D.A., and Vénien-Bryan, C. (2005). Structure 13, this issue, 1463–1472.Google Scholar, clearly shows that there is an intracellular gate formed by the inner helices that move away from the pore in the expanded conformation. The availability of models of KirBac3.1 (Kuo et al., 2003Kuo A. Gulbis J.M. Antcliff J.F. Rahman T. Lowe E.D. Zimmer J. Cuthbertson J. Ashcroft F.M. Ezaki T. Doyle D.A. Science. 2003; 300: 1922-1926Crossref PubMed Scopus (704) Google Scholar) in the closed and open states provides a unique opportunity to examine the importance of microscopic factors affecting channel function using computational methods. The authors have already assessed the stability of a similar open-state model of KirBac1.1 using molecular dynamics simulations (Domene et al., 2005Domene C. Doyle D.A. Venien-Bryan C. Biophys. J. 2005; 89: L01-L03Abstract Full Text Full Text PDF PubMed Scopus (37) Google Scholar) and found, satisfyingly, that it was very stable during extensively long simulations (138 ns). The energetics of ion permeation can also be explored using simple Poisson-Boltzmann (PB) calculations in which the solvent and the membrane are represented as continuum dielectrics. As an illustration, the electrostatic free energy profile of a K+ ion along the pore axis was calculated for the closed and open structures of KirBac1.1 (using the open state model based on KirBac3.1 from Domene et al., 2005Domene C. Doyle D.A. Venien-Bryan C. Biophys. J. 2005; 89: L01-L03Abstract Full Text Full Text PDF PubMed Scopus (37) Google Scholar) and is shown in Figure 1. Clearly, the unfavorable energy barrier at the intracellular bundle crossing in the closed state is completely abolished and, in the open state, the profile is remarkably flat throughout the cytoplasmic domain up to the center of the cavity. These observations support the notion that the latter conformation can conduct ions at a high rate. Figure 1 also shows that a cation in the cavity is electrostatically destabilized in the open state relative to the closed state, a qualitative trend that is observed for many different K+ channels (Jogini and Roux, 2005Jogini, V., and Roux, B. (2005). J. Mol. Biol., in press.Google Scholar). In the future, it is anticipated that the availability of both the closed- and open-state conformations of KirBac3.1 will permit further computational studies that will help resolve current issues about the binding locations of multivalent cations (Mg2+, polyamines), which characteristically cause intracellular blocks of outward ionic flow in inward rectifier channels (John et al., 2004John S.A. Xie L.H. Weiss J.N. Mechanism of inward rectification in Kir channels.J. Gen. Physiol. 2004; 123: 623-625Crossref PubMed Scopus (19) Google Scholar, Shin and Lu, 2005Shin H.G. Lu Z. J. Gen. Physiol. 2005; 125: 413-426Crossref PubMed Scopus (44) Google Scholar). Perhaps the most intriguing aspect of the results of Kuo et al., 2005Kuo, A., Domene, C., Johnson, L.N., Doyle, D.A., and Vénien-Bryan, C. (2005). Structure 13, this issue, 1463–1472.Google Scholar concerns the observation that the presence of Mg2+ stabilizes the closed conformation of the channel in crystallization assays. Electrophysiological studies of Kir channels have long considered the intracellular block by Mg2+ and polyamines to occur through rapid open-state binding from the intracellular side. The location and specific binding of Mg2+ is not fully known, although there are multiple negatively charged residues along the channel pore that have been shown to be involved in binding and current blockade. The results from the work of Kuo et al., 2005Kuo, A., Domene, C., Johnson, L.N., Doyle, D.A., and Vénien-Bryan, C. (2005). Structure 13, this issue, 1463–1472.Google Scholar suggest that the conformation of KirBac3.1 could be directly affected by the presence of Mg2+. The thought that Mg2+ binding could produce such large structural changes is surprising, but it is consistent with a previous study that suggested that spermine blockade in Kir2.1 could induce channel closing (Chang et al., 2003Chang H.K. Yeh S.H. Shieh R.C. J. Physiol. 2003; 553: 101-112Crossref PubMed Scopus (18) Google Scholar). Further investigation will be required to clarify whether the binding of Mg2+ triggers such a conformational change in the channels in their native environment, and, if so, by what mechanism.