Lecture 18 Selectivity Electrophysiology continued: V-gating

1. Lecture 18 Selectivity Electrophysiology continued: V-gating The speed of spike propagation and the geometry of the axon Role of myelination

2. Why are K+ channels highly selective for K+ and Na+ channels are not always highly selective for Na+? From the Goldman treatment we can see that a sufficient depolarization can be easily achieved with a non-selective cationic channel that passes both Na+ and K+. Indeed, we need to depolarize the membrane to the threshold (- 55 mV) to evoke excitation. High selectivity for K+ is required specifically to re-polarize the cell to its resting state (-70 mV) and keep it there.

3. Potassium channel KcsA (from Streptomyces lividans) Selectivity filter carbonyl oxygens (red) substitute for water and provide high selectivity for potassium

4. In the selectivity filter of KcsA the K+ ion is overcoordinated relative to bulk water where only four molecules constitute the first hydration shell around the ion. It’s because water forms electrostatic H-bonds with the surrounding water which largely outcompete the ion. In contrast, there are no H-binding groups around the selectivity filter, which is dedicated to coordinate the ion only. from Varma and Rempe, BJ 2007

5. K+ ion can be coordinated by water (Hydroxyls), formamide (Carbonyls) and by di-glycine (Bidentate carbonyl ligands). The latter mimics the KcsA binding site and the computed free energy of transfer from water to this biding site is close to zero. This is the condition for the fastest transport in/out/across the channel. from Varma and Rempe, BJ 2007

8. Carbonyl oxygens cannot ‘come in touch’ with the smaller Na+ because they start repelling each other (partial charge ~-0.7e on each)

9. According to Molecular Dynamics, the selectivity filter of KscA is not rigid, but can easily ‘collapse’ on a smaller Na+ ion. However, the resultant energy becomes higher due to repulsion between the carbonyl oxygens (see table on the next slide) NMA = N-methylacetamide, a ligand that has carbonyls From Noskov, Berneshe and Roux, Nature,2004

10. From Noskov, Berneshe and Roux, Nature,2004

11. Crowded with Charge Selectivity Filter O½ Wolfgang Nonner, Robert Eisenberg The selectivity filter of L type Ca Channel consists of four Glutamic acid sidechains (EEEE) crowded in a narrow space. Ions are attracted or excluded based on the charge/volume ratio + ++ “E Side Chains”

12. Selective Binding Curve: at approximately 10-6 M Ca2+ displaces Na+ in the selectivity filter L type Ca channel L type Ca Channel Wolfgang Nonner

13. Snap Shots of Contents Radial Crowding is Severe ‘Side Chains’are SpheresFree to move inside channel 6Å In order to convert the channel from Ca to Na-selective, the EEEE motif can be changed to DEKA. All you need is to put the charges into a 6Ǻ confinement Parameters are Fixed in all calculations in all solutions for all mutants Experiments and Calculations done at pH 8 13 Boda, Nonner, Valisko, Henderson, Eisenberg & Gillespie

14. + + The Voltage Sensor + voltage-gated channel o c helices S1-S6 electrometer Voltage dependence of open probability Charged transmembrane helix = voltage sensor

15. Upward motion of voltage-sensor helices (S4)

16. 1 0.8 Po1 ( x ) 0.6 Po2 ( x ) Po3 ( x ) 0.4 0.2 0 0 0 0.05 0.1 0.15 Df, V intrinsic bias o c Variations of the charge in the sensor change both the midpoint and the slope of activation curves

17. 1 0.1 Po1 ( x ) 0.01 Po2 ( x ) 3 Po3 ( x ) . 1 10 4 . 1 10 5 . 1 10 0 0.05 0.1 0.15 Df, V A semi-log plot provides limiting slope for Po at low potentials, which is proportional to z

18. Typical z values for ion channels: Shaker (delayed rectifier) z ~ 13 (3.25/subunit) Various TRP channels z ~ 0.6-2 VDAC (mitochondrial anion channel) z~ 3-4

19. VDAC = voltage dependent anion channel, conducts ATP and ADP Scheme of VDAC gating under positive or negative membrane potentials positive ‘lip’

20. Modification of the gate by polyelectrolytes (dextran sulfate) dramatically changes apparent z in VDAC

21. Properties of different fibers (axons) in the peripheral NS Fiber type Myelination Function Diameter Conduction velocity μm m/s Aa + motoneurons 12-20 70-120 Ab + Touch sensation 5-12 30-70 Ag + muscle spindle 3-6 15-30 Ad no pain, temperature 2-5 12-30 B + visceral afferents, 1-3 3-15 auton. preganglion. C no pain, temperature, 0.3-1.3 0.7-2.5 auton. postganglion.

22. What defines the speed of spike propagation? unmyelinated fiber myelinated fiber =regular wire =High-frequency cable

23. K+ Na+ Na+ K+

24. V1 V2 V2 V1 t time Charging time: t = RC (delay)

25. r L Na+ Internal conductance is proportional to the cross-section: G ~ pr2 R = 1/G, therefore R ~ 1/r2 Capacitance is proportional to the surface area of the cylinder A ~ 2pr L therefore C ~ r Delay t=RC is proportional to 1/r, therefore velocity V ~ r

26. Invertebrates Vertebrates 0.8 mm in diameter 2-5 micrometers

31. Po – intermembrane adhesion protein (immunoglobulin-like) PMP22 – helps compacting the membranes MBP – basic protein remaining in the cytosol Gap junction proteins perforate membranes to allow nutrients

32. From Hille, 2001

33. Myelin coat reduces CAPACITANCE Saltatory excitation in myelinated fibers Fig. 8-17 Velocity is proportional to the internodal distance

34. Fig. 8-13