Sequence Assignments

1
Sequence Assignments
Secondary Structure
Tertiary Structure
2
Secondary Structure
Secondary structure from Chemical
shifts Dipolar couplings J-couplings and
torsion angles Measuring NOEs Measuring
J-couplings
3
Chemical shifts (1H, 13C, 15N) are sensitive to f
and y torsion angles
Cb
H
Ca
N
C
H
H
O
H
N
4
Chemical shifts (1H, 13C, 15N) are sensitive to f
and y torsion angles
CH3
Ca
N
C
H
H
O
Helix Sheet CO 175 ppm 172 ppm Ca 52
ppm 48 ppm Cb 15 ppm 20 ppm
For alanine
5
Secondary structure from short 1H-1H distances
Cb
H
O
dbN
Cb
H
N
C
Ca
N
C
Ca
N
H
H
H
H
H
O
dNN
daN
6
Secondary structure from short 1H-1H distances
Cb
H
O
dbN
Cb
H
N
C
Ca
N
C
Ca
N
H
H
H
H
H
O
dNN Intense NOEs for helix lt 3.6
Å for a-helix
daN Intense NOEs for b-strand
lt 2.7 Å for b-strand
7
Secondary structure from J-couplings
3JHNa
Cb
H
Ca
N
C
H
H
O
3JHNa 6.4 cos2q - 1.4 cosq 1.9 with q f
- 60
b
10
8
a
3JHNa
6
4
2
-160 -120 -80 -40 0
f
8
Secondary structure from J-couplings
3JHNa
Cb
H
Ca
N
C
H
H
O

• 3JHNa 6.4 cos2q - 1.4 cosq 1.9
• -helix (f -57), 3JHNa 3.9 Hz
• 310helix (f -60), 3JHNa 4.2 Hz
• Antiparallel b sheet, (f -139), 3JHNa 8.9 Hz
• Parallel b sheet, (f -119), 3JHNa 9.7 Hz

9
Secondary Structure
Sequence MALRRVETTYGDAWCSTQNLIVWRSTERLN dN
N daN daN(i,i3) 3JHNa
helix
sheet
sheet
3JHNa gt 7 Hz
10
Measuring NOE Intensities
Cb
H
O
dbN
Cb
H
N
C
Ca
N
C
Ca
N
H
H
H
H
H
O
Cb
H
O
dbN
Cb
H
N
C
Ca
N
C
Ca
N
H
H
H
H
H
O
NOE Intensity Distance Weak 3.0 - 5.0
Å Medium 2.0 - 4.0 Å Strong 2.0 - 2.5 Å
11
Measuring J-Couplings
H
O
Glycine
Simplest Spin System - AX
C
N
C
D
H
J
Phase-sensitive COSY
aH
aH
12
Measuring J-Couplings
H
Alanine
H
C
H
O
Spin System - A3X
C
N
C
D
H
H
C
H
C
H
H
H
CH3
Phase-sensitive COSY
H
C
H
H
H
C
H
aH
H
H
C
H
13
Measuring J-Couplings
Spin System - AMX
Serine
A
OD
bH1
H
C
H
O
C
N
C
M
D
H
bH2
Phase-sensitive COSY
X
aH
14
Measuring J-Couplings
Spin System - AMX
Serine
OD
C
Ha
Hm
O
C
C
N
D
Hx
active coupling
Ha
Hm
C
passive coupling
C
Ha
C
Hm
C
C
Hx
Hx
15
Measuring J-Couplings
H
Alanine
H
C
H
O
Spin System - A3X
C
N
C
D
H
active coupling
CH3
Phase-sensitive COSY
passive coupling
aH
Treat only one H of CH3 as active pair
16
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17
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18
Heteronuclear Selective Population Transfer
4
4
C
13
3
3
H
2
1
2
H
1
Saturation of
1,3 transition
C
13
1
1
3,4
1,3
1,3
2,4
1,2
2,4
1,2
3,4
H ..................
C
1
13
19
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20
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21
Carrier between a and b
a on resonance