Determination of the packing mode of the coiled-coil domain of cGMP-dependent protein kinase Ialpha in solution using charge-predicted dipolar couplings.
Coiled-coil motifs are ubiquitous in biology and play essential roles in protein assembly and molecular recognition. Here, we show that the relative orientation and stoichiometry of coiled-coil proteins in solution can be determined by comparison of residual dipolar couplings (RDCs) measured in charged liquid-crystalline medium with values predicted from the three-dimensional charge distribution of the protein. Comparison of charge-predicted RDCs with a small set of one-bond 1DNH dipolar couplings, measured in the negatively charged liquid-crystalline Pf1 bacteriophage medium, identified the coiled-coil region of the cGMP-dependent protein kinase I as a parallel homodimer in solution and ruled out an antiparallel dimeric or monomeric state. The method is very rapid, applicable to a wide variety of liquid crystals used in biological NMR to date, and can be applied to coiled-coil structures and other proteins with higher order assembly.
Zweckstetter
Markus
M
Max Planck Institute for Biophysical Chemistry, Am Fassberg 11, 37077 Göttingen, Germany.
Schnell
Jason R
JR
Chou
James J
JJ
D001665:Binding Sites; D062965:Cyclic GMP-Dependent Protein Kinase Type I; D017869:Cyclic GMP-Dependent Protein Kinases / Q000737:chemistry* / Q000378:metabolism; D019281:Dimerization; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D016012:Poisson Distribution; D012996:Solutions / Q000737:chemistry
D012996:Solutions; D062965:Cyclic GMP-Dependent Protein Kinase Type I; D017869:Cyclic GMP-Dependent Protein Kinases