NMR vs. Crystallography for CHEM 645

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NMR vs. Crystallographyfor CHEM 645
Brian Bahnson Department of Chemistry
Biochemistry University of Delaware
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Distance Restraints
Through space - NOE
NOE ? 1/r6. f (tc)
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Tortional Restraints through bond J-coupling
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NMR Refinement
ideal geometry
NMR term
? wNMR ? distance restraint ? tortional
restraint wideal Etotal
violations violations
X-ray Refinement
ideal geometry
X-ray term
? wF ? whkl (Fo - Fc)2hkl wideal Etotal
hkl
calculated
observed
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13C, 15N labeling, homogeneity
Bigger magnet is better 600, 750 or 900 MHz
2-D and 3-D homonuclear and heteronuclear pulse
sequences
Ikura et al., (1989) Biochemistry 29, 4659-4667.,
then do side chains
NOE Wuthrich, (1989) Science 243, 45-50. also
J-coupling tortion
Pattern recognition, build 100 models, select 20
best
Minimize restraint violations, keep good
geometry ? WN(distance restraint violation
WI (Etotal)
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NMR Structures of closed form calmodulin
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X-ray Crystal Structures of calmodulin
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Bundle of 20 NMR models of calmodulin

• Cases of Bundle Spread
• Missing restraints
• dynamics

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Crystallography vs. NMR advantage/disadvantages

• Experimental difficulties
• need for homogeneity in common
• need good crystals for crystallography
• need 13C and 15N label for NMR
• size limits of NMR technique
• solubility an issue for each technique
• Reported structure(s) look different i.e.
  bundle
• crystal vs. solution structure
• Complementary information
• high resolution vs. dynamics
• positional amplitude,
• certainty time domains

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Molecular Replacement homology modeling
Molecular Replacement (MR) another method to
estimate phases use a structurally homologous
protein gt25 sequence identity is sometimes
possible gt50 sequence identity is a safe bet

• Make search model
• - find structural model of sequence homolog
• - from sequence alignment and homolog structure,
  create model
• - mutate or trim down to what the two proteins
  have in common
• - energy minimize to eliminate bad geometry
  (intro to refinement)

Suppose you wanted to make a model of BSIDH
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Homology Modeling Links
Swiss Model http//swissmodel.expasy.org//SWISS-MO
DEL.html NCBI PubMed http//www.ncbi.nlm.nih.gov/
sites/entrez/ Homology modeling
tutorial http//molvis.sdsc.edu/protexpl/homolmod.
htm Principles of Protein Structure, Comparative
Protein Modeling and Visualization
(http//swissmodel.expasy.org//course/course-inde
x.htm)DeepView    -  download a free version of
this viewer.  Its also for linux
computers.  (http//au.expasy.org/spdbv/) A
tutorial for Deep View was made by Gale Rhodes,
the author of CMCC. (http//www.usm.maine.edu/rh
odes/SPVTut/index.html) 
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