IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1....
Transcript of IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1....
![Page 1: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/1.jpg)
IUPAB NMR Workshop 2009
Exploration of the protein world: Challenges and NMR solutions
R. V. Hosur
Department of Chemical SciencesTata Institute of Fundamental Research
Mumbai 400 005
MBU, June 20, 2008
![Page 2: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/2.jpg)
IUPAB NMR Workshop 2009
C. M. Dobson, Nature 426, 884-890 (2003)
The Protein World: A Structural Biology perspective
The proteome contains a
large number of intrinsically
unfolded proteins
![Page 3: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/3.jpg)
IUPAB NMR Workshop 2009
Triple resonance experiments
![Page 4: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/4.jpg)
IUPAB NMR Workshop 2009
F2 (Cα)
F3 (HN)
i+1i-1 i
F2 (Cα)
F1 (N)
HNCA
F3 (HN)F3 (HN) F3 (HN)
ii
i+1
i-1i-1
i-2
![Page 5: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/5.jpg)
IUPAB NMR Workshop 2009
F2 (Cα)
F3 (HN)
i+1i-1 i
F2 (Cα)
F1 (N)
F3 (HN)F3 (HN) F3 (HN)
HN(CO)CA
i-2
i-1i
![Page 6: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/6.jpg)
IUPAB NMR Workshop 2009
Structure Calculation • Distance Restraints
• Energy Function E = Ef + ENOE
i i il uj j jr r r< <
2 2 - + - ( ) ( )l ul u
NOE ij ij mn mnij mnij mn
k kE r r r r= ∑ ∑
= 0; if the restraint is satisfied
![Page 7: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/7.jpg)
IUPAB NMR Workshop 2009
Restrained Molecular Dynamics Simulation
Distance Geometry
![Page 8: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/8.jpg)
IUPAB NMR Workshop 2009
Protein FoldingLandscape Perspective
Idealized funnel landscape Rugged energy landscape Unfolded ensemble
Intermediate ensemble
Native ensemble
How to monitor the continuous conformational changes as the protein folds ?
![Page 9: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/9.jpg)
IUPAB NMR Workshop 2009
Challenges
Dynamics in proteins
Unfolded and partially folded proteins
Protein folding
Large protein assemblies
![Page 10: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/10.jpg)
IUPAB NMR Workshop 2009
Energy LandscapeEnergy
Denaturant stabilized
![Page 11: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/11.jpg)
IUPAB NMR Workshop 2009
Folded HIV-Pr Unfolded HIV-Pr
![Page 12: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/12.jpg)
IUPAB NMR Workshop 2009
F3 (HN)
F1(N)
F2(N)
3D spectrum with two 15N axes
H
N
2D3D
![Page 13: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/13.jpg)
IUPAB NMR Workshop 2009
An Efficient High-Throughput Resonance Assignment Procedure for StructuralGenomics and Protein Folding Research by NMR
Neel S. Bhavesh, Sanjay C. Panchal, and R. V. Hosur*Biochemistry 40, 14727-14735, 2001.
Accelerated Publications
Improved 3D triple resonance experiments, HNN and HN(C)N, for HNand 15N sequential correlations in (13C, 15N) labeled proteins: Application
to unfolded proteinsSanjay C. Panchal, Neel S. Bhavesh & Ramakrishna V. Hosur∗
Journal of Biomolecular NMR, 20: 135–147, 2001.
A novel protocol based on HN(C)N for rapid resonance assignmentin (15N, 13C) labeled proteins: implications to structural genomicsq
Amarnath Chatterjee, Neel S. Bhavesh, Sanjay C. Panchal,and Ramakrishna V. Hosur,*
Biochemical and Biophysical Research Communications 293 (2002) 427–432
![Page 14: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/14.jpg)
IUPAB NMR Workshop 2009
C N C C N C C N C
O
O O
C
C
CH
H H
α
α
α
β
β
β
i-1 i i+1
HNN
t1
t3t3
t2t2
t2
t3
![Page 15: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/15.jpg)
IUPAB NMR Workshop 2009
F3
F1
F2
NH
15N
15N
HNN
![Page 16: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/16.jpg)
IUPAB NMR Workshop 2009
F3
F1
F2
i
i -1
i +1
i
NH
F1
15N
![Page 17: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/17.jpg)
IUPAB NMR Workshop 2009
F3
F1
F2
i
i -1
i +1
i
i
NH
F2F1
15N 15N
NH
Triplet filter through HSQC
![Page 18: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/18.jpg)
IUPAB NMR Workshop 2009
C N C C N C C N C
O
O O
C
C
CH
H H
αα
α
β
β
β
i-1 i i+1
HN(C)N
t1t2t2
t3
t3
![Page 19: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/19.jpg)
IUPAB NMR Workshop 2009
F2 (N)
F1(N)
F3 (HN)
i-1
ii-1 i
i+1
i
F2 (N)
F1(N)
F3 (HN)i
HN(C)N
![Page 20: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/20.jpg)
IUPAB NMR Workshop 2009
IV
HN
NH
N(C
)N
Special Peak patterns in HNN and HN(C)N
![Page 21: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/21.jpg)
IUPAB NMR Workshop 2009
HNN planes
![Page 22: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/22.jpg)
IUPAB NMR Workshop 2009
Sequential walk through HNN spectrum.
(G) (X) (Z)F 1 (
15N
)
F3 (HN)
G
X
G
X
Z
X
Z
B
-PGXZB-
Neel S. Bhavesh, Sanjay C. Panchal and Ramakrishna V. Hosur. Biochemistry 40, 14727-14735, (2001). (Accelerated publication)
![Page 23: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/23.jpg)
IUPAB NMR Workshop 2009
Alanine Check Points in HNN and HN(C)N SpectraAmarnath Chatterjee, Ashutosh Kumar and Ramakrishna V. Hosur*
J. Magn. Reson. 181, 21-28 (2006)
Tuning the HNN Experiment: Generation of Serine-Threonine Check pointsJeetender Chugh, Dinesh Kumar and Ramakrishna V. Hosur*
J. Biomol. NMR, 40, 145-152 (2008)
![Page 24: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/24.jpg)
IUPAB NMR Workshop 2009
Tuning the HNN
![Page 25: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/25.jpg)
IUPAB NMR Workshop 2009
G63 G63
A11 A11A21 A21
F62 F62
F64 F64
N10 N10
D12 D12
V22 V22
D20 D20
Chatterjee et al, J. Magn. Reson. 181, 21-28 (2006)
Alanine residues behave like glycines in HNN
![Page 26: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/26.jpg)
IUPAB NMR Workshop 2009
Chugh et al J. Biomol. NMR, 40, 145-152 (2008)
![Page 27: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/27.jpg)
IUPAB NMR Workshop 2009
1. HIV-1 protease : 22 kD1. HIV-1 protease, unfolded : 22 kD2. HIV-1 PR precursor : 18 kD (intrinsically unfolded)3. HIV-1 PRTD precursor : 29 kD 4. FK 506 binding protein (FKBP) : 12 kD1. Small ubiquitin like modifier (SUMO): 10 kD1. Small ubiquitin like modifier (SUMO), unfolded : 10 kD2. Drosophila SUMO (folded, unfolded)3. Barstar, unfolded : 10 kD4. Barstar, aggregated (molten globule at low pH) : 160 kD1. GTPase Effector Domain (GED):15 kD, oligomerises (>5
Mda)1. DLC8: dyenin light chain, 10 kD folded2. DLC8 unfolded
Protein systems investigated
![Page 28: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/28.jpg)
IUPAB NMR Workshop 2009
![Page 29: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/29.jpg)
IUPAB NMR Workshop 2009
HIV-I Protease
• 22 kDa Homodimer
• 99 amino acids each
• Helps in maturation of progeny viruses
• Target for anti AIDS drug using protease inhibitor
Flaps
PDB id 1GSL
Active Site
Dimerization domain
Hinge
![Page 30: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/30.jpg)
IUPAB NMR Workshop 2009
Unfolded HIV-I Protease
![Page 31: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/31.jpg)
IUPAB NMR Workshop 2009
Structural propensities
• Secondary Chemical shifts:
δs = δobs – δrc
Cα Hα CO δs + - + α− helix
- + - β- sheet
![Page 32: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/32.jpg)
IUPAB NMR Workshop 2009
Secondary chemical shifts
Neel S. Bhavesh, Sanjay C. Panchal, Rohit Mittal and Ramakrisna V. Hosur. FEBS Lett. 509, 218-24, 2001.
![Page 33: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/33.jpg)
IUPAB NMR Workshop 2009
Residual structural propensities derived from the NMR data are displayed on the crystal structure of HIV-1 protease tethered dimer.
β-structure
α-helixNeel S. Bhavesh, Sanjay C. Panchal, Rohit Mittal and Ramakrisna V. Hosur. FEBS Lett. 509, 218-24, 2001.
Local structural preferences: Initial folding events
![Page 34: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/34.jpg)
IUPAB NMR Workshop 2009
6M Gdn 5M Gdn
![Page 35: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/35.jpg)
IUPAB NMR Workshop 2009
![Page 36: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/36.jpg)
IUPAB NMR Workshop 2009
Folding propensities in HIV-1 protease
∆R2 = 3.0 – 4.5 s-1 ∆R2 = 1.5 – 3.0 s-1
![Page 37: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/37.jpg)
IUPAB NMR Workshop 2009
SUMO
Mishra, et al, J. Biol. Chem. 279, 31445-31454 (2004)
![Page 38: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/38.jpg)
IUPAB NMR Workshop 2009
![Page 39: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/39.jpg)
IUPAB NMR Workshop 2009
Ashutosh Kumar et al, Biophys. J. 90, 2498-2509 (2006)Ashutosh Kumar et al, J. Mol. Biol. 361, 180-194 (2006)
Equilibrium populations down the funnel
Secondary shifts
8M urea
7M urea
6M urea
5M urea
4M urea
3M urea
![Page 40: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/40.jpg)
IUPAB NMR Workshop 2009
Line broadening : micro-milli second time scale conformational transitions
Ashutosh Kumar et al, J. Mol. Biol. 361, 180-194 (2006)
![Page 41: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/41.jpg)
IUPAB NMR Workshop 2009
N
N
C
1M0M
8MN
C
N
C
7M 6MN
C
5M
C
N
4M
C
3M
C
N
C
N
Ashutosh Kumar et al, J. Mol. Biol. 361, 180-194 (2006)
Equilibrium populations down the funnel
![Page 42: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/42.jpg)
IUPAB NMR Workshop 2009
NMR Insights into Protein Function
![Page 43: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/43.jpg)
IUPAB NMR Workshop 2009
Random Coil
Pre molten Globule
Molten Globule
Folded
Structure Function Paradigm
Random Coil
Pre-Molten Globule
Molten Globule
Folded
AggregatedUversky V. N. (2002) Prot. Sci. 11, 739
![Page 44: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/44.jpg)
IUPAB NMR Workshop 2009
Protein Dynamics
Tumbling motions
Internal high frequency motions (nano- to pico- second time scales)
Conformational transitions (milli- to micro- second time scales)
Functional significance
![Page 45: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/45.jpg)
IUPAB NMR Workshop 2009
Active site
HIV-1 protease
Structure – Dynamics - Function
![Page 46: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/46.jpg)
IUPAB NMR Workshop 2009
Heavy Heavy ChainsChains
Intermediate Intermediate ChainsChains
DLC8DLC8
Dynein Light Chain Protein (DLC8)
DLC8DLC8 – – DimerDimerpH 7pH 7
DyneinDynein
MicrotubuleMicrotubule
CargoCargoMoleculeMolecule
![Page 47: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/47.jpg)
IUPAB NMR Workshop 2009
Dynamic regions
Dynamics: R2, Curved temp dependence, Line broadening
![Page 48: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/48.jpg)
IUPAB NMR Workshop 2009
Dynamics, pH 7
Dynamics, pH 6
![Page 49: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/49.jpg)
IUPAB NMR Workshop 2009
![Page 50: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/50.jpg)
IUPAB NMR Workshop 2009DLC8 binding with (VYTKQTQTTST)
Free protein, pH 7
Spectrum at pH 7,DLC8-peptide complex
Spectrum at pH 6,Free protein + complex
Recognition motif(K/R)XTQT
pH Switch for cargo trafficking
![Page 51: IUPAB NMR Workshop 2009 Exploration of the protein world: …iupab/WS_Nov26_2009_4.pdf · 2010. 1. 4. · IUPAB NMR Workshop 2009 An Efficient High-Throughput Resonance Assignment](https://reader035.fdocuments.net/reader035/viewer/2022071214/604358b2724ea339eb3a568b/html5/thumbnails/51.jpg)
IUPAB NMR Workshop 2009
Ph. D. Students
Sanjay C. PanchalNeel S. BhaveshAmarnath ChatterjeeAshutosh KumarJeetendra ChughP. M. KrishnamohanManoj RoutDinesh KumarSwagata Chakraborty
Post-docs
Ragini SinhaManeesha BarveShilpi Sharma
Others
Dr. Rohit MittalJyoti Ranjan MishraRam Kumar MishraAnindya Ghosh Roy
Dr. M.V. HosurDr. B. Pillai