Chapter 3 : Deciphering KcsA as a K Channel Model

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As a member of the family Streptomycetaceae, is a gram-positive soil bacterium with a complex growth cycle. In the presence of nutrients this growth is initiated by the germination of spores. Giant liposome-protoplast vesicles derived from the mutant strain carrying the plasmid pKCS1 (with the K channel of streptomycetes A (KcsA) gene) showed ion channel activity under neutral pH (7.2) and asymmetric conditions. The negatively charged lipid phosphatidylglycerol (PG) and the nonbilayer lipid PE support tetramerization and membrane association of KcsA better than the zwitterionic bilayer lipid phosphatidylcholine (PC). In vitro studies using a transcription-translation system revealed that no tetramer is formed in a membrane-free reaction but only after the addition of inner membrane vesicles. With a coupled in vitro transcription-translation system, highest tetramerization was recorded in the presence of pure lipid vesicles, demonstrating that a phospholipid bilayer is the minimal requirement to form the KcsA tetramer. Polyhydroxybutyrate (PHB) and inorganic phosphate (polyP) are widely distributed among prokaryotic and eukaryotic organisms. The results of experimental and comparative structural data support the conclusion that the crystal structure of the tetrameric KcsA does not present the open state. The majority of the virus progeny is released about 8 h after infection. The current model assumes that Kcv is located in the internal membrane of the virus and will be inserted into the plasma membrane of the host cell. The hydration state of K ions and their permeation need to be reinvestigated.

Citation: Schrempf H. 2005. Deciphering KcsA as a K Channel Model, p 41-68. In Kubalski A, Martinac B (ed), Bacterial Ion Channels and Their Eukaryotic Homologs. ASM Press, Washington, DC. doi: 10.1128/9781555816452.ch3

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Amino Acids
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Figure 1

Features of and the generation of giant protoplasts. (A) Vegetative network of hyphae. (B) Protoplasts of were obtained by digesting (with lysozyme) the cell wall of the hyphae. (C) General scheme to fuse protoplasts with liposomes to generate giant protoplast vesicles. (D) Current recording at pH 7.2 under asymmetric conditions and the deduced histogram to present the proportion of closed and open channels (top). Current recordings and all point histograms after the addition of CsCl (bottom). (E) Current recordings (excised patch) under asymmetric conditions within ranging potentials.

Citation: Schrempf H. 2005. Deciphering KcsA as a K Channel Model, p 41-68. In Kubalski A, Martinac B (ed), Bacterial Ion Channels and Their Eukaryotic Homologs. ASM Press, Washington, DC. doi: 10.1128/9781555816452.ch3
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Image of Figure 2
Figure 2

Characteristics of the deduced and the isolated KcsA protein as well as electrophysiological analysis of using a bilayer. (A) The characteristic regions of the KcsA protein deduced from the corresponding gene. (B) The KcsA protein assembles to a stable tetramer (left), which upon increase of temperature, can be resolved via intermediates (middle; above 70°C) to the monomeric form (right; melting point close to 85°C). (C) Generation of proteoliposomes and subsequent fusion with a bilayer. (D) Current recordings of a bilayer containing KcsA at pH 7.2 under asymmetric conditions. (E) Current recordings at different combinations of pH at and sites under symmetric conditions.

Citation: Schrempf H. 2005. Deciphering KcsA as a K Channel Model, p 41-68. In Kubalski A, Martinac B (ed), Bacterial Ion Channels and Their Eukaryotic Homologs. ASM Press, Washington, DC. doi: 10.1128/9781555816452.ch3
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Figure 3

Comparative electrophysiological characteristics of KcsA wild-type (WT) and mutant proteins generated after exchange of residues L81 and Y82. (A) The relative positioning of the relevant amino acids within KcsA. (B) Relative open probabilities of the channels (white bars, −100 mV; black bars, +100 mV). (C) Comparative sensitivity to TEA of WT and mutant channels (white bars, −100 mV; black bars, +100 mV). (D) Voltage dependence of WT and mutant channels of external TEA block. (E) Rectification properties of different proteins.

Citation: Schrempf H. 2005. Deciphering KcsA as a K Channel Model, p 41-68. In Kubalski A, Martinac B (ed), Bacterial Ion Channels and Their Eukaryotic Homologs. ASM Press, Washington, DC. doi: 10.1128/9781555816452.ch3
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1. Alvis, S. J.,, I. M. Williamson,, J. M. East,, and A. G. Lee. 2003. Interactions of anionic phospholipids and phosphatidylethanolamine with the potassium channel KcsA. Biophys. J. 85:38283838.
2. Caprini, M.,, S. Ferroni,, R. Planells-Cases,, J. Rueda,, C. Rapisarda,, A. Ferrer-Montiel,, and M. Montal. 2001. Structural compatibility between the putative voltage sensor of voltage-gated K+channels and the prokaryotic KcsA channel. J. Biol. Chem. 276:2107021076.
3. Chen, G. Q.,, C. Cui,, M. L. Mayer,, and E. Gouaux. 1999. Functional characterization of a potassium-selective prokaryotic glutamate receptor. Nature 402:817821.
4. Choe, S.,, A. Kreusch,, and P. J. Pfaffinger. 1999. Towards the three-dimensional structure of voltage-gated potassium channels. Trends Biochem. Sci. 24:345349.
5. Compoint, M.,, P. Carloni,, C. Ramseyer,, and C. Girardet. 2004. Molecular dynamics study of the KcsA channel at 2.0-A resolution: stability and concerted motions within the pore. Biochim. Biophys. Acta 1661:2639.
6. Coronado, R.,, R. L. Rosenberg,, and C. Miller. 1980. Ionic selectivity, saturation, and block in a K+-selective channel from sarcoplasmic reticulum. J. Gen. Physiol. 76:425446.
7. Cortes, D. M.,, L. G. Cuello,, and E. Perozo. 2001. Molecular architecture of full-length KcsA. Role of cytoplasmic domains in ion permeation and activation gating. J. Gen. Physiol. 117:165180.
8. Criado, M.,, and B. U. Keller. 1987. A membrane fusion strategy for single-channel recordings of membranes usually non-accessible to patch-clamp pipette electrodes. FEBS Lett. 224:172176.
9. Cuello, L. G.,, J. G. Romero,, D. M. Cortes,, and E. Perozo. 1998. pH-dependent gating in the Streptomyces lividans K+channel. Biochemistry 37:32293236.
10. Demmers, J. A.,, A. van Dalen,, B. de Kruijff,, A. J. Heck,, and J. A. Killian. 2003. Interaction of the K+channel KcsA with membrane phospholipids as studied by ESI mass spectrometry. FEBS Lett. 541:2832.
11. Derst, C.,, and A. Karschin. 1998. Evolutionary link between prokaryotic and eukaryotic K+channels. J. Exp. Biol. 201:27912799.
12. Doyle, D. A.,, J. M. Cabral,, R. A. Pfuetzner,, A. Kuo,, J. M. Gulbis,, S. L. Cohen,, B. T. Chait,, and R. MacKinnon. 1998. The structure of the potassium channel: molecular basis of K+conduction and selectivity. Science 280:6977.
13. Durell, S. R.,, and H. R. Guy. 2001. A family of putative Kir potassium channels in prokaryotes. BMC Evol. Biol. 1:14.
14. Durell, S. R.,, Y. Hao,, T. Nakamura,, E. P. Bakker,, and H. R. Guy. 1999. Evolutionary relationship between K(+) channels and symporters. Biophys. J. 77:775788.
15. Enkvetchakul, D.,, J. Bhattacharyya,, I. Jeliazkova,, D. K. Groesbeck,, C. A. Cukras,, and C. G. Nichols. 2004. Functional characterization of a prokaryotic Kir channel. J. Biol. Chem. 279:4707647080.
16. Enz, C.,, T. Steinkamp,, and R. Wagner. 1993. Ion channels in the thylakoid membrane (a patch clamp study). Biochim. Biophys. Acta 1143:76.
17. Fink, M.,, F. Duprat,, F. Lesage,, R. Reyes,, G. Romey,, C. Heurteaux,, and M. Lazdunski. 1996. Cloning, functional expression and brain localization of a novel unconventional outward rectifier K+channel. EMBO J. 15:68546862.
18. Fox, J. A. 1987. Ion channel subconductance states. J. Membr. Biol. 97:18.
19. Grupe, A.,, K. H. Schroeter,, J. P. Ruppersberg,, M. Stocker,, T. Drewes,, S. Beckh,, and O. Pongs. 1990. Cloning and expression of a human voltage-gated potassium channel. A novel member of the RCK potassium channel family. EMBO J. 9:17491756.
20. Guidoni, L.,, V. Torre,, and P. Carloni. 1999. Potassium and sodium binding to the outer mouth of the K+channel. Biochemistry 38:85998604.
21. Gulbis, J. M.,, and D. A. Doyle. 2004. Potassium channel structures: do they conform? Curr. Opin. Struct. Biol. 14:440446.
22. Heginbotham, L.,, L. Kolmakova-Partensky,, and C. Miller. 1998. Functional reconstitution of a prokaryotic K+channel. J. Gen. Physiol 111:741749.
23. Heginbotham, L.,, M. LeMasurier,, L. Kolmakova-Partensky,, and C. Miller. 1999. Single Streptomyces lividans K+channels. Functional asymmetries and sidedness of proton activation. J. Gen. Physiol. 114:551559.
24. Heginbotham, L.,, Z. Lu,, T. Abramson,, and R. MacKinnon. 1994. Mutations in the K+channel signature sequence. Biophys. J. 66:10611067.
25. Hellmer, J.,, and C. Zeilinger. 2003. MjK1, a K+channel from M. jannaschii, mediates K+uptake and K+sensitivity in E. coli. FEBS Lett. 547:165169.
26. Hille, B. 1992. Ionic Channels of Excitable Membranes. Sinauer Associates Inc., Sunderland, Mass.
27. Holyoake, J.,, C. Domene,, J. N. Bright,, and M. S. Sansom. 2003. KcsA closed and open: modelling and simulation studies. Eur. Biophys. J. 33:238246.
28. Inagaki, N.,, T. Gonoi,, J. P. I. Clement,, N. Namba,, J. Inazawa,, G. Gonzalez,, L. Aguilar-Bryan,, S. Seino,, and J. Bryan. 1995. Reconstitution of IKATP: an inward rectifier subunit plus the sulfonylurea receptor. Science 270:11661170.
29. Jiang, Y.,, A. Lee,, J. Chen,, V. Ruta,, M. Cadene,, B. T. Chait,, and R. MacKinnon. 2003. X-ray structure of a voltage-dependent K+channel. Nature 423:3341.
30. Kessler, A.,, W. Dittrich,, M. Betzler,, and H. Schrempf. 1989. Cloning and analysis of a deletable tetracyclineresistance determinant of Streptomyces lividans 1326. Mol. Microbiol. 3:11031109.
31. Kuo, M. M.,, Y. Saimi,, and C. Kung. 2003. Gain-of-function mutations indicate that Escherichia coli Kch forms a functional K conduit in vivo. EMBO J. 15:40494058.
32. Kutzner, H. J., 1981. The family Streptomycetaceae, pp. 20282090. In M. P. Starr,, H. Stolp,, H. G. Trüper,, A. Balows,, and H. Schlegel (ed.), The Prokaryotes: a Handbook on Habitats, Isolation and Identification of Bacteria. Springer-Verlag, Berlin, Germany.
33. Lauger, P. 1985. Structural fluctuations and current noise of ionic channels. Biophys. J. 48:369373.
34. Le Dain, A. C.,, P. J. Anderton,, D. K. Martin,, and T. J. Millar. 1994. A tetraethylammonium-insensitive inward rectifier K+channel in Muller cells of the turtle (Pseudemys scripta elegans) retina. J. Membr. Biol. 141:239245.
35. Legros, C.,, V. Pollmann,, H. G. Knaus,, A. M. Farrell,, H. Darbon,, P. E. Bougis,, M. F. Martin-Eauclaire,, and O. Pongs. 2000. Generating a high affinity scorpion toxin receptor in KcsA-Kv1.3 chimeric potassium channels. J. Biol. Chem. 275:1691816924.
36. LeMasurier, M.,, L. Heginbotham,, and C. Miller. 2001. KcsA: it’s a potassium channel. J. Gen. Physiol. 118: 303314.
37. Lu, Z.,, A. M. Klem,, and Y. Ramu. 2002. Coupling between voltage sensors and activation gate in voltage-gated K+channels. J. Gen. Physiol. 120:663676.
38. Luneau, C.,, R. Wiedmann,, J. S. Smith,, and J. B. Williams. 1991. Shaw-like rat brain potassium channel cDNA’s with divergent 3′ends. FEBS Lett. 288:163167.
39. Luzhkov, V. B.,, F. Osterberg,, and J. Aqvist. 2003. Structure-activity relationship for extracellular block of K+ channels by tetraalkylammonium ions. FEBS Lett. 554:159164.
40. Mehmel, M.,, M. Rothermel,, T. Meckel,, J. L. Van Etten,, A. Moroni,, and G. Thiel. 2003. Possible function for virus encoded K+channel Kcv in the replication of chlorella virus PBCV-1. FEBS Lett. 552:711.
41. Meuser, D.,, H. Splitt,, R. Wagner,, and H. Schrempf. 1999. Exploring the open pore of the K+channel KcsA from Streptomyces lividans. FEBS Lett. 462:447452.
42. Meuser, D.,, H. Splitt,, R. Wagner,, and H. Schrempf. 2001. Mutations stabilizing an open conformation within the external region of the permeation pathway of the potassium channel KcsA. Eur. Biophys. J. 30:385391.
43. Milkman, R. 1994. An Escherichia coli homologue of eucaryotic potassium channel proteins. Proc. Natl. Acad. Sci. USA 91:35103514.
44. Mueller, P.,, D. O. Rudin,, H. Tien,, and W. C. Wescott. 1962. Reconstitution of cell membrane structure in vitro and its transformation into an excitable system. Nature (London) 194:979980.
45. Navarro, B.,, M. E. Kennedy,, B. Velimirovic,, D. Bhat,, A. S. Peterson,, and D. E. Clapham. 1996. Nonselective and Gβγ-insensitive weaver K+channels. Science 272:19501953.
46. Nelson, P. H. 2003. Modeling the concentration-dependent permeation modes of the KcsA potassium ion channel. Phys. Rev. E. 68:061908.
47. Pascual, J. M.,, C.-C. Shieh,, G. E. Kirsch,, and A. M. Brown. 1995. Multiple residues specify external tetraethylammonium blockade in voltage-gated potassium channels. Biophys. J. 69:428434.
48. Patlak, B. J. 1988. Sodium channel subconductance levels measured with a new variance-mean analysis. J. Gen. Physiol. 98:413430.
49. Perozo, E.,, D. M. Cortes,, and L. G. Cuello. 1998. Three-dimensional architecture and gating mechanism of a K+channel studied by EPR spectroscopy. Nature Struct. Biol. 5:459469.
50. Perozo, E.,, D. M. Cortes,, and L. G. Cuello. 1999. Structural rearrangements underlying K+-channel activation gating. Science 285:7378.
51. Reusch, R. N. 1999. Streptomyces lividans potassium channel contains poly-(R)-3-hydroxybutyrate and inorganic polyphosphate. Biochemistry 38:1566615672.
52. Schrempf, H., 1999. Investigations of streptomycetes using tools of recombinant DNA technology, p. 501510. In A. L. Demain, and J. E. Davies (ed.), Manual of Industrial Microbiology and Biotechnology, 2nd ed. ASM Press, Washington, D.C.
53. Schrempf, H.,, O. Schmidt,, R. Kümmerlen,, S. Hinnah,, D. Müller,, M. Betzler,, T. Steinkamp,, and R. Wagner. 1995. A prokaryotic potassium ion channel with two predicted transmembrane segments from Streptomyces lividans. EMBO J. 14:51705178.
54. Schwarz, T. L.,, B. L. Tempel,, D. M. Papazian,, Y. N. Jan,, and L. Y. Jan. 1988. Multiple potassium-channel components are produced by alternative splicing at the Shaker locus in Drosophila. Nature 331:137142.
55. Slesinger, P. A.,, N. Patil,, Y. J. Liao,, Y. N. Jan,, L. Y. Jan,, and D. R. Cox. 1996. Functional effects of the mouse weaver mutation on G protein-gated inwardly rectifying K+channels. Neuron 16:321331.
56. Splitt, H.,, D. Meuser,, I. Borovok,, M. Betzler,, and H. Schrempf. 2000. Pore mutations affecting tetrameric assembly and functioning of the potassium channel KcsA from Streptomyces lividans. FEBS Lett. 472:8387.
57. Trudeau, M. C.,, J. W. Warmke,, B. Ganetzky,, and G. A. Robertson. 1995. HERG, a human inward rectifier in the voltage-gated potassium channel family. Science 269:9295.
58. Valiyaveetil, F. I.,, Y. Zhou,, and R. MacKinnon. 2002. Lipids in the structure, folding, and function of the KcsA K+channel. Biochemistry 41:1077110777.
59. van Dalen, A.,, S. Hegger,, J. A. Killian,, and B. de Kruijff. 2002a. Influence of lipids on membrane assembly and stability of the potassium channel KcsA. FEBS Lett. 525:3338.
60. van Dalen, A.,, H. Schrempf,, J. A. Killian,, and B. de Kruijff. 2000. Efficient membrane assembly of the KcsA potassium channel in Escherichia coli requires the protonmotive force. EMBO Rep. 1:340346.
61. van Dalen, A.,, L. M. van der Laan,, A. J. Driessen,, J. A. Killian,, and B. de Kruijff. 2002b. Components required for membrane assembly of newly synthesized K+channel KcsA. FEBS Lett. 511:5158.
62. van den Brink-van der Laan, E.,, V. Chupin,, J. A. Killian,, and B. de Kruijff. 2004. Small alcohols destabilize the KcsA tetramer via their effect on the membrane lateral pressure. Biochemistry 43:59375942.
63. Warmke, J.,, R. Drysdale,, and B. Ganetzky. 1991. A distinct potassium channel polypeptide encoded by the Drosophila eag locus. Science 252:15601562.
64. Wolters, M.,, M. Madeja,, A. M. Farrell,, and O. Pongs. 1999. Bacillus stearothermophilus lctB gene gives rise to functional K+channels in Escherichia coli and in Xenopus oocytes. Receptors Channels 6:477491.
65. Woodhull, A. M. 1973. Ionic blockage of sodium channels in nerve. J. Gen. Physiol. 61:687708.
66. Yool, A. J.,, and T. L. Schwarz. 1991. Alteration of ionic selectivity of a K+channel by mutation of the H5 region. Nature 349:700704.
67. Zakharian, E.,, and R. N. Reusch. 2004a. Functional evidence for a supramolecular structure for the Streptomyces lividans potassium channel KcsA. Biochem. Biophys. Res. Commun. 322:10591065.
68. Zakharian, E.,, and R. N. Reusch. 2004b. Streptomyces lividans potassium channel KcsA is regulated by the potassium electrochemical gradient. Biochem. Biophys. Res. Commun. 316:429436.
69. Zhou, Y.,, J. H. Morais-Cabral,, A. Kaufman,, and R. MacKinnon. 2001. Chemistry of ion coordination and hydration revealed by a K+channel-Fab complex at 2.0 A resolution. Nature 414:4348.

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