Simultaneous detection and deconvolution of congested NMR spectra containing three isotopically labeled species.
We present a procedure for isolating subspectra corresponding to individual protein or peptide components in a ternary mixture or complex. Each of the three-component species is labeled differently: species A uniformly with 15N, species B uniformly with 15N and 13C, and species C uniformly with 15N but selectively with 13C' or 13Calpha. By using the dual carbon label selective HSQC (DCLS-HSQC) pulse sequence and exploiting differences in 1J 15N-13C coupling patterns to filter selected 15N resonances from detection during a constant time period, a subspectrum from each species can be generated from three spectra acquired from a single sample. Many important biological pathways involve dynamic interactions among members of multicomponent protein assemblies, and this approach offers a powerful way to monitor such processes.
Department of Chemistry and Biochemistry, University of Minnesota, Minneapolis, Minnesota 55455-0431, USA.
Larry R
LR
Masterson
Marco
M
Tonelli
John L
JL
Markley
Gianluigi
G
Veglia
D002247:Carbon Isotopes / Q000737:chemistry; D006859:Hydrogen / Q000737:chemistry; D007553:Isotope Labeling / Q000379:methods*; D009587:Nitrogen Isotopes / Q000737:chemistry; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods*; D011506:Proteins / Q000032:analysis* / Q000737:chemistry
D002247:Carbon Isotopes; D009587:Nitrogen Isotopes; D011506:Proteins; D006859:Hydrogen