Optimal methyl labeling for studies of supra-molecular systems

• Published in: Journal of biomolecular NMR 2010-07, Vol.47 (3), p.163-169
• Main Authors: Religa, Tomasz L, Kay, Lewis E
• Format: Article
• Language: English
• Subjects: Biochemistry; Biological and Medical Physics; Biophysics; Carbon Isotopes; Deuterium; HMQC; HSQC; Isoleucine; Leucine; Methyl labeling; Methyl-TROSY; Methylation; Nuclear Magnetic Resonance, Biomolecular - methods; Physics; Physics and Astronomy; Probes; proteasome endopeptidase complex; Proteasome Endopeptidase Complex - chemistry; Proteins - chemistry; Sensitivity; Spectroscopy; Spectroscopy/Spectrometry; Valine
• ISSN: 0925-2738; 1573-5001
• DOI: 10.1007/s10858-010-9419-7

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.