bcmb / chem 8190 biomolecular nmr graduate course...
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BCMB / CHEM 8190 Biomolecular NMRGRADUATE COURSE OFFERING IN NUCLEAR
MAGNETIC RESONANCE
"Biomolecular Nuclear Magnetic Resonance" is a course intended for all graduate students with an interest in applications of nuclear magnetic resonance (NMR) to problems in structural and functional biology. It will begin with a treatment of the fundamentals that underlie magnetic resonance phenomena and develop this into a basis for experimental design, interpretation of data, and critical reading of the literature.
http://tesla.ccrc.uga.edu/courses/bionmr/
SyllabusI. Introduction
A. M 1/9 Prestegard Magnetic properties of nuclei and electrons – precession 5-38 L*
B. W 1/11 Prestegard RF pulses and spin relaxation -Bloch equations 39-50 L*, 653 L*
II. InstrumentationA. M 1/16 MLK Jr. Holiday (no class)B. W 1/18 Prestegard Instrumental considerations -
a look at probes 65-76 L*C. M 1/23 Prestegard Fourier transform methods
and data Processing 85-102 L*
• Friday Labs are scheduled separately (C122) – intro to computer systems 1/13, 1/20
• Texts: "Spin Dynamics - Basics of Nuclear Magnetic Resonance", M. H. Levitt (L), “Protein NMR Spectroscopy, Principles & Practice“, J. Cavanagh, W. J. Fairbrother, A. G. Palmer III, N. J. Skelton. (C)
Biomolecular NMR 2012• Biomolecular NMR - short history ~ 1985 first
protein structure• Compared to X-ray ~ 1953 first protein
structure• Today ~ 13 % of structures in the PDB come
via NMR – higher for nucleic acids• Unique structural applications – weak
associations, partially structured, membrane associations, in-cell observation
• Diverse applications: drug screening, metabolic monitoring, in vivo imaging
• NMR is still an evolving science
Unstructured Regions in Proteins% with rmsd > 4 Å
0
2
4
6
8
10
12
14
16
18
0 0.1 0.2 0.3 0.4 0.5 0.6
Protein fraction
Freq
uenc
y
NMRXray
Can NMR Make a Unique Contribution ?Membrane Proteins, Protein-Protein Interactions, Ligand Binding
• 1952 - Bloch and Purcell – Nobel Prize in Physics• 1991 – Richard Ernst – Nobel Prize in Chemistry• 2002 – Kurt Wuthrich – Nobel Prize in Chemistry• 2003 – Lauterbur and Mansfield – Nobel Prize in
Physiology and Medicine
NMR Recognition
Computers Were Primitive in 1966
Varian HR 220
Early Superconducting
Magnets Required a Lot of
Care And Feeding
High Field (220 MHz), but Still 1D CW NMR
80’s Approach: 2D 1H-1H NMR: 9 kDa Proteinassign resonances, collect NOE’s, calculate structure
- - - C N C C N - - -α
Hα
Cβ
= =- -
O O
H
H H
HH
- - - C N C C N - - -α
Hα
Cβ
= =
- -
O O
H
H H
HH
(HB)CBCA(CO)NH
HNCACB
Extension to 3D: Through-bond Correlations in PeptidesIsotope Labeling is Key
IKURA M; KAY LE; BAX A, (1990) BIOCHEMISTRY 29:4659-4667
1D2D
3D
f3 (1H)
f1 (13C)
f2 (15N)
HN(CO)CA
Today Very Large Systems Can Be Studied:Proteasome subunit – active site dynamics
Sprangers, R and Kay, LE, 2007. Probing supramolecular structure from measurement of methyl H-1-C-13 residual dipolar couplings. Journal of the American Chemical Society 129: 12668-+.
Ruschak AM and Kay LE, 2010, Methyl groups as probes of supra-molecular structure, dynamics and function, J Biomol NMR 46:75-87
NMR is widely applicable to structure and function of biomolecules
• Judge, P. J. & Watts, A. (2011). Recent contributions from solid-state NMR to the understanding of membrane protein structure and function. Current Opinion in Chemical Biology 15, 690-695.
• Kang, C. B. & Li, Q. X. (2011). Solution NMR study of integral membrane proteins. Current Opinion in Chemical Biology 15, 560-569.
• Dominguez, C., Schubert, M., Duss, O., Ravindranathan, S. & Allain, F. H. T. (2011). Structure determination and dynamics of protein-RNA complexes by NMR spectroscopy. Progress in Nuclear Magnetic Resonance Spectroscopy 58, 1-61.
• Montelione, G. T. & Szyperski, T. (2010). Advances in protein NMR provided by the NIGMS Protein Structure Initiative: Impact on drug discovery. Current Opinion in Drug Discovery & Development 13, 335-349.
• Pacheco-Torres, J., Lopez-Larrubia, P., Ballesteros, P. & Cerdan, S. (2011). Imaging tumor hypoxia by Magnetic Resonance methods. Nmr in Biomedicine 24, 1-16.
• Powers, R. (2009). NMR metabolomics and drug discovery. Magnetic Resonance in Chemistry 47, S2-S11.
• Tycko, R. (2011). Solid-State NMR Studies of Amyloid Fibril Structure. In Annual Review of Physical Chemistry, Vol 62 (Leone, S. R., Cremer, P. S., Groves, J. T. & Johnson, M. A., eds.), Vol. 62, pp. 279-299.
• Tzeng, S. R. & Kalodimos, C. G. (2011). Protein dynamics and allostery: an NMR view. Current Opinion in Structural Biology 21, 62-67.
Spin PropertiesB0
μ = γ I Iz = hm E = -μ⋅B0
B0
m = -1/2
m = 1/2
ν = γB0/2π Mz = γhΣipimiN0
= N0γ h B0 I(I+1)
3kT
22
p = exp(-Ei/kT)/Z
for I = 1/2
E
NMR Active Isotopes Exist for Nearly Every Element
http://bouman.chem.georgetown.edu/NMRpt/NMRPerTab.html
Select an element by clicking on it:
1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 H H He Li Be B C N O F Ne Na Mg Al Si P S Cl Ar K Ca Sc Ti V Cr Mn Fe Co Ni Cu Zn Ga Ge As Se Br Kr Rb Sr Y Zr Nb Mo Tc Ru Rh Pd Ag Cd In Sn Sb Te I Xe Cs Ba * Hf Ta W Re Os Ir Pt Au Hg Tl Pb Bi Po At Rn Fr Ra ** Rf Ha Sg Ns Hs Mt *La Ce Pr Nd Pm Sm Eu Gd Tb Dy Ho Er Tm Yb Lu **Ac Th Pa U Np Pu Am Cm Bk Cf Es Fm Md No Lr
Isotope: 13C
Spin: 1/2 Natural abunance: 1.108% Magnetogyric ratio (rad/T s): 6.7283 x 10^7 Relative receptivity: 1.76 x 10^-4 Magnetic moment 1.2166 Quadrupole moment Q/m(2) 0 Resonance frequency 25.144 MHz
Spin ½ Nuclei are Most Useful in Biomolecualr NMR
Polypeptides are Rich in NMR Active Nuclei
Me – 1H
Amide 1HCα 1H
Amide 15N
Cβ 13C
C’ 13C Cα 13C
Nuclear Properties
• Not all nuclei have magnetic moments, Why?• Not all nuclei are equally abundant, Why?• Spins vary, Why?• Magnetogyric ratios vary, Why?
Fundamental Particle Properties
N
S
Stern Gerlach experiment: Na atom - 1 unpaired electronTwo spots implies quantized moments: +/- 1/2protons and neutrons are also spin 1/2 particles
e- Beam in fieldgradient
Current Loop Model for Magnetic MomentsM
iArea, S
M = i S i in Coulomb s-1
S in m2
M in JT-1 (Tesla)
Estimates: i = -ev/(2πr), S = πr2, M (or μ) = -erv/2
μ = -e(r x v)/2, L = me r x v, μ = -e/(2me) L = γ L = γh/(2π)l
γ = -g (e/(2me)), g = Lande g factor
r
Values of Particle Magnetogyric Ratios
Electron: g ≈ 2, γe = -17.7 x 1010 T-1s-1
Proton: expect 1/mp dependence, 1/2000 and positive2.7 x 108 T-1s-1
Neutron: similar mass to proton-1.8 x 108 T-1s-1
Heavier Nuclei: the Shell Model
Simple Rules:
a) spherical particle in a box potentialψ = Rnl(r) Yl
m(θ,φ), E(n,l)ladder of energy levels like H atom, but all ls allowed
b) strong coupling of spin and orbit angular momentumquantized total: j = l +/- 1/2 for spin 1/2 particlelarger j, lower energy (usually)
V(r)
r0r0 0
Shell Model Rules Continued
c) Treat protons and neutrons separately and fill from bottom up assuming 2j + 1 degeneracy
d) Assume particle pair strongly within levels: only unpaired spins count - total spin angular momentum given by j of level for unpaired spin
e) sign of moment depends on sign of moment for fundamental particle but changes sign when moment subtracts instead of adds to l in giving j
Energy Level Diagram
n+1 j degeneracy total
2s 1/2 2 20
3/2 41d
5/2 6
1/2 2 81p
3/2 4
1s 1/2 2 2
Examples of Nuclear Properties
• 168O - most abundant in Earth’s crust. 8 protons, 8
neutrons - 2 magic #s. All particles paired - no moment.
• 136C - 6 protons, 7 neutrons. All protons paired. One
neutron unpaired in 1p1/2. j = l - 1/2. Therefore, negative moment subtracts from l. Positive γ spin 1/2 particle