Angular Dependence of 3-bond J-couplings

Bax, et al. (1994) Measurement of Homo- and Heteronuclear J-couplings from Quantitative J Correlation, Methods Enzymol., 239, 79-105

Detection of Hydrogen Bonds

h3JNC’ -0.2 to -0.9

h2JHC’ -0.6 to 1.3

h3JHC 0.0 to 1.4

Ref: Grzesiek, et al. (2001) Methods Enzymol., 338, 111-133

Anisotropic Tumbling w.r.t. to BAnisotropic Tumbling w.r.t. to Boo

Results in Residual Dipolar Couplings (RDCs)Results in Residual Dipolar Couplings (RDCs) • Magnitude of the dipole-

dipole interaction is orientation dependent w.r.t. to the static magnetic field (Bo)

• Isotropic tumbling w.r.t. Bo normally averages dipolar couplings to zero

• Small, but non-zero, magnetic susceptibility results in residual dipolar couplings that appear as apparent J-splittings

Induced Residual Alignment of Diamagnetic Induced Residual Alignment of Diamagnetic ProteinsProteins

• Lipid Bicelles LC (Tjandra & Bax, Science, 1997)• Purified Bacteriophage Particles (Pf1) LC(Hansen et al, J. Am.

Chem. Soc, 1998)• Deformed Pores in Nondenaturing Polyacrylamide Gel (Sass

et al, J. Biomol. NMR, 2000)

dimyristoyl-phos-phatidylcholine (DMPC)

dihexanoyl-phos-phatidylcholine (DHPC)

RDC for Proteins in Solution Correlate Very Well WithPredictions from High-Resolution Crystal Structures

NMR Structure Determination 1. Start with a peptide chain of random starting

conformation

2. Subject protein to a classical mechanical treatment (such as “simulated annealing”) that minimizes the total energy

3. Simulated annealing protocol is a commonly usedto minimize the energy. Protein is heated in the computer,which allows molecular motions to occur, and then is slowly cooled to minimize the energy (avoids local minima in the energy landscape)