Side Chain Interactions In Helices
62
Side chain Side chain Interactions in Interactions in α α -helices -helices Ph.D. Dissertation Defense 04/23/08 Prashant Girinath Dept. of Chemistry SUNY- University at Buffalo
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Transcript of Side Chain Interactions In Helices
Side chainSide chain Interactions in Interactions in αα-helices-helices
Ph.D. Dissertation Defense04/23/08
Prashant GirinathDept. of Chemistry
SUNY- University at Buffalo
H: Alpha HelixS: SheetT: TurnR: Random Coil
-40
-20
0
20
40
60
80
103
·
θ (d
eg·c
m2 ·d
mol
-1 )
180 260220λ (nm)
H
R
S
T
The α-Helix
Pauling, L. et. al. Proc. Natl. Acad. Sci. U. S. A. 1951, 37, 205-207Barlow, D. J.; Thornton, J. M. J. Mol. Biol. 1988, 168, 601-619.
www.aveda.comwww.time.com
(i, i+4)
(i, i+3)
top viewside view
Strop, P.; Mayo, S. L. Biochemistry 2000, 39, 1251-1255.
Lysine (Lys, K)
OH3N
O
NH3
+
+
Arginine (Arg, R)
OH3N
O
HN
+
HN
NH3
+
Aspartic acid (Asp, D)
OH3N
O
+
O-
O
Glutamic acid (Glu, E)
OH3N
O
+
O O
- - - -
-
Charged amino acids
Does Side chain Length Matter?
n=1, Dap, S-2,3-diamonipropionic acid 2, Dab, S-2,4-diaminobutyric acid 3, Orn, L-ornithine 4, Lys, L-lysine
NH
O
H3N
n
+
Peptide Sequence
EXaa3: Ac-YGG A EAAX A EAAX A EAAX A-NH2
EXaa4: Ac-YGG A EAAAX EAAAX EAAAX A-NH2
EXaa5: Ac- YGG EAAAAX EAAAAX EAAAAX -NH2
Xaa (X): Dap, Dab, Orn, Lys
w Lys = 0.93w Orn = 0.63w Dab = 0.32w Dap = 0.14
Baldwin, R. L. et. al J. Mol. Biol. 1996, 257, 726-734Marqusee, S.; Baldwin, R. L. Proc. Natl. Acad. Sci. U. S. A. 1987, 84, 8898-8902
ELys5EOrn5EDab5EDap5
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200 210 220 230 240 250
[!]
(10
3·d
eg
·cm
2·d
mo
l-1)
" (nm)
EXaa5 CD at pH 7
EXaa5 CD pH 2 - pH 12
ELys5EOrn5EDab5EDap5
-30
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-20
-15
-10
-5
0
2 4 6 8 10 12
[!] 2
22 (
10
3·d
eg·c
m2·d
mol-1
)
pH
H: Alpha HelixS: SheetT: TurnR: Random Coil
-40-20
020406080
103
·θ
(deg
·cm2 ·d
mol
-1 )
180 260220λ (nm)
H
R
S
T
EXaa5 CD at pH 2
ELys5EOrn5EDab5EDap5
-40
-20
0
20
40
60
80
200 210 220 230 240 250
[!]
(10
3·d
eg
·cm
2·d
mo
l-1)
" (nm)
ELys4EOrn4EDab4EDap4
-40
-20
0
20
40
60
80
190 200 210 220 230 240 250
[ !] (1
03·d
eg·c
m2·d
mol-1
)
" (nm)
EXaa4 CD at pH 7
EXaa4 CD pH 2 - pH 12
ELys4EOrn4EDab4EDap4
-30
-25
-20
-15
-10
-5
0
2 4 6 8 10 12
[!] 2
22 (
10
3·d
eg
·cm
2·d
mo
l-1)
pH
ELys4EOrn4EDab4EDap4
-40
-20
0
20
40
60
80
190 200 210 220 230 240 250
[ !] (1
03·d
eg·c
m2·d
mol-1
)
" (nm)
EXaa4 CD at pH 2
Scholtz, J. M et.al. Biochemistry 1993, 32, 9668-9676
Lys = robust interaction
Lys - 1 methylene ≠ robustness
ELys3EOrn3EDab3EDap3
-20
-10
0
10
20
30
40
190 200 210 220 230 240 250
[!]
(10
3·d
eg
·cm
2·d
mo
l-1)
" (nm)
EXaa3 CD at pH 7
EXaa3 CD pH 2 - pH 12
ELys3EOrn3EDab3EDap3
-30
-25
-20
-15
-10
-5
0
2 4 6 8 10 12
[ !] 2
22 (
10
3·d
eg
·cm
2·d
mo
l-1)
pH
ELys3EOrn3EDab3EDap3
-20
-10
0
10
20
30
190 200 210 220 230 240 250
[!]
(10
3·d
eg
·cm
2·d
mo
l-1)
" (nm)
EXaa3 CD at pH 2
Lys = interaction
Lys - 1 methylene ≠ interaction
Molecular Modeling
Dunbrack, R. L.; Karplus, M. J. Mol. Biol. 1993, 230, 543-574.
Stability : t > g+ > g-
ELys4:ELys4: (t, g+)(t, g+)((g-, g+g-, g+))
Others: ((t, g+t, g+))(i)
(i+4)
Interacting Species
Lowest Energy Conformations
Cheng, R. P. et. al. Biochemistry 2007, 46, 10528-10537
(i)
(i+3)
Stability : t > g+ > g-
ELys3:ELys3: ((t, tt, t))((g+, g+g+, g+))
Others: ((t, t, g-g-))
Interacting Species
Lowest Energy Conformations
Cheng, R. P. et. al. Biochemistry 2007, 46, 10528-10537
Cheng, R. P. et. al. Biochemistry 2007, 46, 10528-10537
What about Asp?What about Asp?
Lysine (Lys, K)
OH3N
O
NH3
+
+
Arginine (Arg, R)
OH3N
O
HN
+
HN
NH3
+
Aspartic acid (Asp, D)
OH3N
O
+
O-
O
Glutamic acid (Glu, E)
OH3N
O
+
O O
- - - -
-
Glu Glu - One - One methylene methylene = Asp= Asp
NH
O
O-
On
n=1, Asp (D), L-aspartic acid 2, Glu (E), L-glutamic acid 3, Aad (J), S-2-aminoadipic acid
n=1, Dap, S-2,3-diamonipropionic acid 2, Dab, S-2,4-diaminobutyric acid 3, Orn, L-ornithine 4, Lys, L-lysine
NH
O
H3N
n
+
Peptide Sequence
ZXaa3: Ac-YGG A ZAAX A ZAAX A ZAAX A-NH2
ZXaa4: Ac-YGG A ZAAAX ZAAAX ZAAAX A-NH2
ZXaa5: Ac- YGG ZAAAAX ZAAAAX ZAAAAX -NH2
Z: D(Asp), E(Glu), J(Aad)
Xaa (X): Dap, Dab, Orn, Lys
w Aad = 1.19w Glu = 0.45w Asp = 0.31
DXaa4 CD at pH 7
DLys4DOrn4DDab4DDap4
-40
-20
0
20
40
60
200 210 220 230 240 250
[!] (1
03·d
eg·c
m2·d
mol-1
)
" (nm)
DXaa4 CD pH 2 - pH 12
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-5
0
2 4 6 8 10 12
DLys4DOrn4DDab4DDap4
[!] 2
22 (1
03.d
eg
.cm
2.d
mol-1
)
pH
DXaa4 CD at pH 2
DLys4DOrn4DDab4DDap4
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0
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20
30
40
200 210 220 230 240 250
[!]
(10
3·d
eg
·cm
2·d
mo
l-1)
" (nm)
For environment sensitive (i, i+4)interactions
NatureNature uses Asp!uses Asp!
DXaa3 CD at pH 7
DLys3DOrn3DDab3DDap3
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10
20
30
40
200 210 220 230 240 250
[!] (1
03·d
eg·c
m2·d
mol-1
)
" (nm)
DXaa3 CD pH 2 - pH 12
-35
-30
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-10
-5
0
2 4 6 8 10 12
DLys3DOrn3DDab3DDap3
[ !] 2
22 (1
03.d
eg.c
m2.d
mol-1
)
pH
DXaa3 CD at pH 2
DLys3DOrn3DDab3DDap3
-30
-20
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0
10
20
30
40
200 210 220 230 240 250
[ !] (1
03·d
eg·c
m2·d
mol-1
)
" (nm)
Asp = (Glu - 1 methylene)
Asp ≠ robust interaction
For robust interactions
Nature needs Nature needs GluGlu!!
Glu + 1 methylene = Aad = ?
AadLys4AadOrn4AadDab4AadDap4
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200 210 220 230 240 250
[!]
(10
3·d
eg
·cm
2·d
mo
l-1)
" (nm)
AadXaa4 CD at pH 7
AadXaa4 CD pH 2 - pH 12
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-5
0
2 4 6 8 10 12
AadLys4AadOrn4AadDab4AadDap4
[!] 2
22 (1
03.d
eg
.cm
2.d
mo
l-1)
pH
AadLys4AadOrn4AadDab4AadDap4
-40
-20
0
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40
60
200 210 220 230 240 250
[!] (1
03·d
eg·c
m2·d
mol-1
)
" (nm)
AadXaa4 CD at pH 2
AadXaa3 CD at pH 7
AadLys3AadOrn3AadDab3AadDap3
-30
-20
-10
0
10
20
30
40
200 210 220 230 240 250
[ !]
(10
3·d
eg
·cm
2·d
mo
l-1)
" (nm)
AadXaa3 CD pH 2 - pH 12
-35
-30
-25
-20
-15
-10
-5
0
2 4 6 8 10 12
AadLys3AadOrn3AadDab3AadDap3
[!] 2
22 (1
03.d
eg.c
m2.d
mol-1
)
pH
AadXaa3 CD at pH 2
AadLys3AadOrn3AadDab3AadDap3
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-20
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40
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[!]
(10
3·d
eg
·cm
2·d
mo
l-1)
" (nm)
Nature does not need Aad!
1) How robust is robust?
2) Nature + Innovation = ?
NH
O
O-
On
n=1, Asp (D), L-aspartic acid 2, Glu (E), L-glutamic acid 3, Aad (J), S-2-aminoadipic acid
n=1, Dap, S-2,3-diamonipropionic acid 2, Dab, S-2,4-diaminobutyric acid 3, Orn, L-ornithine 4, Lys, L-lysine
NH
O
H3N
n
+
Peptide Sequence
XaaZ3: Ac-YGG A XAAZ A XAAZ A XAAZ A-NH2
XaaZ4: Ac-YGG A XAAAZ XAAAZ XAAAZ A-NH2
XaaZ5: Ac- YGG XAAAAZ XAAAAZ XAAAAZ -NH2
Z: D(Asp), E(Glu), J(Aad)
Xaa (X): Dap, Dab, Orn, Lys
XaaZ4 CD at pH 7
LysAad4OrnAad4DabAad4DapAad4
-30
-20
-10
0
10
20
30
40
200 210 220 230 240 250
[!]
(10
3·d
eg
·cm
2·d
mo
l-1)
" (nm)
LysAsp4OrnAsp4DabAsp4DapAsp4
-30
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20
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40
200 210 220 230 240 250
[!]
(10
3·d
eg
·cm
2·d
mo
l-1)
" (nm)
LysGlu4OrnGlu4DabGlu4DapGlu4
-30
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30
40
200 210 220 230 240 250
[!] (1
03·d
eg·c
m2·d
mol-1
)
" (nm)
XaaZ4 CD at pH 2
LysGlu4OrnGlu4DabGlu4DapGlu4
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[!]
(10
3·d
eg
·cm
2·d
mo
l-1)
" (nm)
LysAsp4OrnAsp4DabAsp4DapAsp4
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-20
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40
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[ !]
(10
3·d
eg
·cm
2·d
mo
l-1)
" (nm)
LysAad4OrnAad4DabAad4DapAad4
-30
-20
-10
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20
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40
200 210 220 230 240 250
[!]
(10
3·d
eg
·cm
2·d
mo
l-1)
" (nm)
XaaZ3 CD at pH 7
LysGlu3OrnGlu3DabGlu3DapGlu3
-30
-20
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30
40
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[!]
(10
3·d
eg
·cm
2·d
mo
l-1)
" (nm)
LysAad3OrnAad3DabAad3DapAad3
-30
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-10
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40
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[ !]
(10
3·d
eg
·cm
2·d
mo
l-1)
" (nm)
LysAsp3OrnAsp3DabAsp3DapAsp3
-30
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0
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40
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[ !]
(10
3·d
eg
·cm
2·d
mo
l-1)
" (nm)
XaaZ3 CD at pH 2
LysGlu3OrnGlu3DabGlu3DapGlu3
-20
-10
0
10
20
30
40
200 210 220 230 240 250
[!]
(10
3·d
eg
·cm
2·d
mo
l-1)
" (nm)
LysAad3OrnAad3DabAad3DapAad3
-20
-10
0
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20
30
40
200 210 220 230 240 250
[ !]
(10
3·d
eg
·cm
2·d
mo
l-1)
" (nm)
LysAsp3OrnAsp3DabAsp3DapAsp3
-20
-10
0
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30
40
200 210 220 230 240 250
[ !]
(10
3·d
eg
·cm
2·d
mo
l-1)
" (nm)
Nature + Human Innovation =
NH
O
Aib
10θ
(deg
·cm
·dm
ol
λ (nm)
2
The 310 helix
Therapeutically relevantfolding intermediate to α-helix
EKEK
EK KE
AA
A
A
E A E K A K E A E K A K
(i,i+3) E-K, K-E stabilizes310 helix
(i,i+4) E-E, K-K destabilizesαHelix
(i,i+2) E-E, K-K destabilizesrandom coil
Ln: Ac-YGG-(EAEKAK)n-NH2
310-Helix Design
-0.01
0
0.01
0.02
0.03
0.04
0.05
0.06
0.07
161016201630164016501660167016801690
A
U
! cm-1
L1
L2
VaLd-A
L4
L5
L6
L3
1655Random coil
1650α-Helix
16351635331010-Helix-Helix
Amide I band νcm-1
Structure
IR Spectra
-3 104
-2.5 104
-2 104
-1.5 104
-1 104
-5000
0
200 210 220 230 240 250
[!]
(deg
cm
2 d
mo
l-1)
" (nm)
L1
L2
L3
L4
L5
L6
CD Spectra
NMR
Ac Y1 G2 G3 E4 E6 E10 E12 K15 E16
d!N(i,i+1)
NH2
d!N(i,i+3)
d!"(i,i+3)
d#N(i,i+1)
A5 K7 A8 K9 A11 K13 A14 A17 E18 K19 K21A20
d#N(i,i+2)
d#N(i,i+3)
d!N(i,i+2)
d!"(i,i+1)
d!"(i,i+2)
NMR inconclusive due to sequence degeneracy
CD and IR indicate presence of 310-helix
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