Potassium channels: permeation, selectivity, inactivation, and gating

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Potassium channels permeation, selectivity,
inactivation, and gating Biological
Membranes Structure and Function Ohio Center
for Technology and Science 8-11 October,
2005 Benoît Roux Department of
Biochemistry Department of Physiology
Biophysics Weill Medical College of Cornell
University benoit.roux_at_med.cornell.edu http//tha
llium.med.cornell.edu/RouxLab
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selectivity filter

gate Channel opens at low intracellular pH
(Cuello et al, 1998 Heginbotham et al, 1999)
X-ray structure of KcsA (Doyle et al, 1998)
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Permeation
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Traditional phenomenologies
FqEmp
Z
Traditional approaches, such as Eyring Rate
Theory or Nernst-Planck continuum
electrodiffusion theory, picture the movements of
ions across membrane channels as chaotic random
displacements in a free energy profile W(z)
driven by the transmembrane electric field
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Molecular Dynamics simulations of the KcsA K
Channel
FMA 150 mM KCl 112 DPPC, 6500 waters Over
40,000 atoms No cutoff of electrostatics
(PME) Nanosecond simulations Bernèche
Roux (Biophys J, 2000)
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Bernèche Roux. Energetics of ion conduction
through the K channel Nature 414, 73 (2001)
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Brownian Dynamics Simulations of K in KcsA
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Elementary Microscopic Step for Ion Conduction
KChannel is a narrow multiion pore with 2-3
ions The knock-on mechanism of Hodgkin
Keynes (1955)
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Internal TEA Blockade
Kutluay, Roux Heginbotham (2004)
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Ion Selectivity
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Ion Conduction selectivity are governed free
energy profiles
FqEmp
Z
K
??G G(Na in channel) G(Na in bulk)
- G(K in channel) G(K in bulk)
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The snug-fit model of ion selectivity
The coordination cage provided by the rigid pore
is such that K fits precisely, but Na is too
small to be well coordinated, so it does not
enter the pore (Bezanilla Armstrong, 1972)
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The paradox of selectivity
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Selectivity for K against Na
-1.12.86.62.4-1.5
-1.762.105.221.07-1.85
0.75 8.24 10.60 5.20 3.18

1K4C 1K4C 1BL8
S0
S1
S2
S3
S4
flexible
rigid
Molecular Dynamics Alchemical Free Energy
Perturbations
Reference in bulk ?G 21.0 kcal/molCV
Reference in bulk ?G 18.0 kcal/molCV
Sergey Noskov, 2004
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FEP calculations with channel in membrane
Including all interactions 5.63
S2
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Simple model of a charge surrounded by carbonyl
dipoles
Free Energy (kcal/mol)
K vs Na selectivity increase with the number
of carbonyls
Noskov, Berneche, Roux Control of ion
selectivity in potassium channels by
electrostatic and dynamic properties of carbonyl
ligands Nature 2004 431830-4
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Inactivation
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Can the selectivity filter adopt other
conformations?
Fast ion conduction
Non-conducting?
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Conducting states of the selectivity filter
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K current under voltage-clamp conditions
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Single channel recordings of wild-type KcsA (Po
0.15)
pH jump Macroscopic current recordings using
liposome patch clamp
Cordero, Cuello, Perozo
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The inactivation gate is located at the
selectivity filter
WT/G116-SL
The inner gate formed by the helix bundle is
unaffected by the same mutation affecting the
open probability
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Crystal structure of E71A mutant Configuration 1
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Crystal structure of E71A mutant Configuration 2
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In Shaker W434F inactivates
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Ion occupancy in the filter
Flipped filter

• In E71A non-flipped structure ion configuration
  is same as WT
• In the flipped structure there are 5 ions in the
  filter and no ion in Sext

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Gating
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Sliding helix
Large movement
Paddle
Small movement
Transporter-like
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Jiang et al (2003)
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S1
S2
S4
S3
S6
S5
X-ray structure of Kv1.2 (pdb id 2A79) Long et
al (Science, 2005)
Model of Shaker Chanda et al (Nature, 2005)
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Crystallographic structure of Kv1.2
Long et al (Nature, 2005)
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Z (Å)
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(from x-ray structure)
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3
1
2
(from MD)
Bulk water
Lipids
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2
1
4
3
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Vmp
i
side I
side II



Cl- Ag ? AgCl e-
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Vmp
i
side I
side II



AgCl
Cl-
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Vmp
i
e-
e-
e-
e-
side I
side II
e-
e-
e-
e-
e-

Cl-
Cl-
AgCl
Cl-
Cl-
Cl-
Cl-
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Microscopic Theory of Gating Charge
Solve numerically the PB-V equation for the open
and the closed states to calculated the total
gating charge
B. Roux, The influence of the membrane potential
on the free energy of an intrinsic protein,
Biophys. J. 73, 2980 (1997).
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Chanda, Asamoah, Blunck, Roux and Bezanilla.
Nature 436, 852-856 (2005) Gating charge
displacement in voltage-gated ion channels
involves limited transmembrane movement.
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Acknowledgements Sergey Noskov, Simon Bernèche
(selectivity) Vishwanath Jogini (MD of
Kv1.2) Yanxiang Zhao, Vishwanath Jogini (X-ray
structures) Eduardo Perozo, Julio Cordero, Luis
Cuello, Sudha Chakrapani Francisco Bezanilla,
Baron Chanda,, Osei Kwame Asamoah, Rikard
Blunck Olaf Andersen, Toby Allen, George
Eisenman NIH, NSF, NCSA PSC
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carbonyls 8 7 6 5 4 6 5 4
water 0 1 2 3 4 0 1 2
??G (kcal/mol) 6.20 4.79
2.28 -0.69 -2.11 3.40 3.19 0.26
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Microscopic Theory of Membrane Potential
-

-

-

PB-V
B. Roux, The influence of the membrane potential
on the free energy of an intrinsic protein,
Biophys. J. 73, 2980 (1997).
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There are 7 favorable (binding) sites for K

Bernèche Roux, (2001)