Optimal methyl labeling for studies of supra-molecular systems
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Selective methyl labeling combined with HMQC spectroscopy that exploits a TROSY effect in 13CH3 spin systems has significantly extended the utility of solution NMR spectroscopy in studies of high molecular weight particles. Herein we compare the utility of 13CH3- versus 13CHD2-labeling of Ile, Leu, Val probes in supra-molecular systems through quantification of relative signal-to-noise ratios in optimized spectra of highly deuterated, 13CH3- and 13CHD2-labeled samples of the half proteasome (α7α7, 360 kDa). It is shown that the sensitivity of spectra recorded on Ile, Leu, Val 13CH3-labeled samples is between 1.5 and 2 fold higher than the corresponding data sets obtained on α7α7 with 13CHD2 probes. Thus, labeling of supra-molecules with 13CH3 isotopomers remains the method of choice, but in applications where 13CHD2 moieties are required, sensitivity will in general not be limiting.
KeywordsMethyl-TROSY Methyl labeling Sensitivity HSQC HMQC Proteasome
T.L.R. acknowledges The European Molecular Biology Organization (ALTF 827-2006) and The Canadian Institutes of Health Research (CIHR) for postdoctoral fellowships. L.E.K. holds a Canada Research Chair in Biochemistry. This work was supported by a grant from the CIHR.
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