http://proteinsandwavefunctions.blogspot.com/2011/02/drug-design-and-ph.html

1. Drug Design and pH!
Jan H. Jensen!
Department of Chemistry!
University of Copenhagen!
http://propka.ki.ku.dk/~jhjensen!
1

2. Drug Design and pH!
Getting the protonation state right
on the ligand
and the protein!
The protonation state determines
!
Charge and hydrogen bonding properties
!
2

3. 3

4. pKa value => charge as a function of pH!
1
q( pH ) =
1 + 10 pH − pKa
4

5. The protonation state determines charge
and hydrogen bonding properties !
1ACJ.pdb!
5

6. Determining pKa values for ligands!
Experimental:!
Spectroscopic feature vs pH!
NMR Shift!
pH!
Computational:
!
substituents
Hammet-Taft rules pK a = pK model + ρ ! ∑ σi
i
PH+  P + H+ pK a = ΔGrxn!/ RT ln(10)
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7. Ligand pKa prediction software
using Hammet-Taft rules!
Epik
!
http://www.schrodinger.com/products/14/4/
!
Marvin!
http://www.chemaxon.com/marvin/sketch/index.jsp!
MoKa!
http://www.moldiscovery.com/soft_moka.php
!
ACD/pKa DB!
http://www.acdlabs.com/products/phys_chem_lab/pka/
!
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8. Drug Design and pH!
Getting the protonation state right
on the ligand 
and the protein !
The protonation state determines
!
Charge and hydrogen bonding
properties
!
8

9. Standard pKa values for residues !
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10. Non standard pKa values for residues in proteins !
Picture of non-standard!
Protonation state in active site!
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11. Simple rules for pKa Predictions?!
AH  A- + H+!
pKa = 4.5!
pKa = ΔGrxn/1.36!
pKa = 4.8 - 0.3!
pKa = 4.8 - 0.5 + 0.2!
pKa = 5.0!
pKa = 4.8 + 0.2!
pKa = 4.8!
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12. PropKa!
pK a = pK a + ΔpK a
model Desolv
+ ΔpK a + ΔpK a
HB Chg-Chg
Model pKa values:
C-End=3.20, Asp=3.80, Glu=4.50, His=6.50, Cys=9.00, Tyr=10.00,
N-End=8.0, Lys=10.50, Arg=12.50.
PROPKA1: Li, Robertson & Jensen Proteins 2005!
(PROPKA3: Olsson, Rostkowski, Søndergaard, Jensen JCTC 2011)! 12

13. pKa: Hydrogen Bonding!
! !∆pKa= -0.8 , if D < d1!
∆pKa= -0.8  (D-d2) / (d1-d2), if D < d2!
! !!
∆pKa
! !∆pKa= 0.0, if D > d2!
O
F0
O H O
D
0.0
0.0 d1 d2 D (Å)
Li, Robertson & Jensen Proteins 2005! 13

14. Example: Asp102 in RNase H1!
N 15.5Å=443
∆pKa= +0.43 Arg46
Asp148
-1.20
15.5 Å +0.73 Arg46
Asp102
4.5 Å -1.20
-0.46 -2.40
-0.48 Asp102
Leu103
NLocal =13
a ∆pKa= +0.91
b c
pKa = 3.8 + 1.3 - 3.3 - 1.7 = +0.1!
Exp = < 2.0
Li, Robertson & Jensen Proteins 2005! 14

15. Ca 20,000 hits last 12 months!
Included in PDB2PQR and Vega-ZZ (*)! 15

16. 16

17. PROPKA-VMD interface!
17
Rostkowski, Olsson, Søndergaard, Jensen BMC Struct. Biol. 2011!

18. 12 14
(a) Asp /Glu (b) Cys
10
12
8
PROPKA Prediction
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PROPKA Prediction
6
8
4
6
2
0 RMSD = 0.7! 4
RMSD = 1.0!
-2
N = 210! 2 N = 11!
-2 0 2 4 6 8 10 12 2 4 6 8 10 12 14
Experimental pKa values Experimental pKa values
10 14
(c) His (d) Lys
12
8
PROPKA Prediction
10
PROPKA Prediction
6
8
4
6
2
4
RMSD = 1.2! RMSD = 0.7!
0 N = 41! 2
N = 24! 18
0 2 4 6 8 10 2 4 6 8 10 12 14
Experimental pKa values Experimental pKa values

19. pKa values can change upon ligand binding!
Kb0
P+L P·L
pK a − pH
c
H+ H+ 1 + 10
Ka f
Ka c
K obs = K 0
b pK af − pH
PH+ + L PH+·L
1 + 10
Kb+
⎛ [PH + iL] ⎞ ⎛ [H + ] ⎞
⎜ 1 + [PiL] ⎟
[PiL] ⎝ ⎠ ⎜1 + K c ⎟
0 ⎝ a ⎠
+
[PiL] + [PH iL]
K obs = = = Kb
[P][L] + [PH + ][L] [P][L] ⎛ [PH + ] ⎞ ⎛ [H + ] ⎞
⎜ 1 + [P] ⎟
⎝ ⎠ ⎜1 + K f ⎟
⎝ a ⎠
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20. pKa values can change upon ligand binding!
Implication number 1:!
Inhibition constant is pH dependent!
pK a − pH
c
0 1 + 10
K obs = K b
1 + 10 pKa − pH
f
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21. pKa values can change upon ligand binding!
Implication number 2:!
Change in Ki wrt pH means !
change in protonation stateupon binding!
∂ log(K obs )
− = qc − q f
q=
1 ∂pH
1 + 10 pH − pKa
qf
K obs = K 0
b
qc 21

22. pKa values can change upon ligand binding!
Implication number 3:!
Docking score using static
protonation state must be corrected!
⎛ 1+ 10 pK ac − pH ⎞
ΔGb = −RT ln(K b ) − RT ln⎜
0
pK − pH
f ⎟ = ΔGb + ΔGb, pH
0 0
⎝ 1+ 10 a ⎠
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23. pKa values can change upon ligand binding!
Implication number 4:!
ΔH measured by calorimetry will be
buffer dependent and must be corrected!
ΔH corrected = ΔH obs − (qc − q f )ΔH ion
ΔHion is ionization enthalpy of buffer! 23

24. Effect of Ligands: PROPKA 2.0!
pK a = pK a + ΔpK a
model Desolv
+ ΔpK a + ΔpK a
HB Chg-Chg
Atom typing!
H-bond donor/acceptor!
Charged groups!
Ligand ionizable groups/pKmodel!
PROPKA 2: Bas, Rogers & Jensen Proteins 2008!
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PROPKA3.1: Søndergaard & Jensen, in progress!

25. xxx.pdb!
PROPKA!
xxx.pka new_xxx.pdb!
edit!
new_xxx.pdb!
PROPKA!
new_xxx.pka!
pKmodel!
Edit = !
new pKmodel or!
new atom types!
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26. 1K1L.pdb -> 1K1L.pka!
pKa! pKmodel!
10.3!
12.0!
3.2!
Experiment!
new_1K1L.pdb!
N29!
new_1K1L.pka! C91! C25!
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27. 27
Bas, Rogers & Jensen Proteins 2008!

28. Summary!
Implications for docking!
Protonation state of ligand can be estimated
computationally!
Protonation states of active site residues are not!
always “standard”!
Protonation states can change upon binding!
In which case docking score must be corrected!
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29. Questions Now?!
Questions Later?!
Leave a comment on!
http://proteinsandwavefunctions.blogspot.com/2011/02/drug-design-and-ph.html!
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