Characterization of specific protein association by 15N CPMG relaxation dispersion NMR: the GB1(A34F) monomer-dimer equilibrium.
The Carr-Purcell-Meiboom-Gill (CPMG) transverse relaxation dispersion NMR experiment is a powerful means for detecting and characterizing conformational exchange. This experiment reports the exchange of chemical shifts and therefore can monitor all chemical exchange phenomena, not only intramolecular conformational exchange. Here, we report a CPMG transverse relaxation dispersion study for the monomer-dimer equilibrium of the GB1 point mutant, Ala-34-Phe (GB1(A34F)). This variant exists predominantly as a side-by-side dimer at high concentration (>1 mM). We demonstrate that the dispersion experiment is exceptionally valuable for studying association equilibria since it is extremely sensitive to the minor population in the equilibrium. Twenty-eight individual amide sites in the GB1(A34F) dimer protein were monitored via a 2D (15)N-(1)H HSQC spectroscopy, and all relaxation-derived data are consistent with predominantly an exchange process between dimer and monomer species.
Jee
Jungoo
J
Department of Structural Biology, University of Pittsburgh School of Medicine, Biomedical Science Tower 3, 3501 Fifth Avenue, Pittsburgh, Pennsylvania 15260, USA.
Ishima
Rieko
R
Gronenborn
Angela M
AM
D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D019281:Dimerization; D008958:Models, Molecular; D009587:Nitrogen Isotopes; D019906:Nuclear Magnetic Resonance, Biomolecular; D017434:Protein Structure, Tertiary; D013291:Streptococcus / Q000737:chemistry / Q000235:genetics / Q000378:metabolism
D001426:Bacterial Proteins; D009587:Nitrogen Isotopes