Dataset Information

Nitrogen-detected TROSY yields comparable sensitivity to proton-detected TROSY for non-deuterated, large proteins under physiological salt conditions.

ABSTRACT: Direct detection of the TROSY component of proton-attached (15)N nuclei ((15)N-detected TROSY) yields high quality spectra with high field magnets, by taking advantage of the slow (15)N transverse relaxation. The slow transverse relaxation and narrow line width of the (15)N-detected TROSY resonances are expected to compensate for the inherently low (15)N sensitivity. However, the sensitivity of (15)N-detected TROSY in a previous report was one-order of magnitude lower than in the conventional (1)H-detected version. This could be due to the fact that the previous experiments were performed at low salt (0-50 mM), which is advantageous for (1)H-detected experiments. Here, we show that the sensitivity gap between (15)N and (1)H becomes marginal for a non-deuterated, large protein (? c = 35 ns) at a physiological salt concentration (200 mM). This effect is due to the high salt tolerance of the (15)N-detected TROSY. Together with the previously reported benefits of the (15)N-detected TROSY, our results provide further support for the significance of this experiment for structural studies of macromolecules when using high field magnets near and above 1 GHz.

SUBMITTER: Takeuchi K

PROVIDER: S-EPMC4871712 | biostudies-literature | 2016 Feb

REPOSITORIES: biostudies-literature

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Nitrogen-detected TROSY yields comparable sensitivity to proton-detected TROSY for non-deuterated, large proteins under physiological salt conditions.

Takeuchi Koh K Arthanari Haribabu H Imai Misaki M Wagner Gerhard G Shimada Ichio I

Journal of biomolecular NMR 20160122 2

Direct detection of the TROSY component of proton-attached (15)N nuclei ((15)N-detected TROSY) yields high quality spectra with high field magnets, by taking advantage of the slow (15)N transverse relaxation. The slow transverse relaxation and narrow line width of the (15)N-detected TROSY resonances are expected to compensate for the inherently low (15)N sensitivity. However, the sensitivity of (15)N-detected TROSY in a previous report was one-order of magnitude lower than in the conventional (1 ...[more]

PMID: 26800993

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Project description:The discovery of the TROSY effect (Pervushin et al. in Proc Natl Acad Sci USA 94:12366-12371, 1997) for reducing transverse relaxation and line sharpening through selecting pathways in which dipole-dipole and CSA Hamiltonians partially cancel each other had a tremendous impact on solution NMR studies of macromolecules. Together with the methyl TROSY (Tugarinov and Kay in J Biomol NMR 28:165-172, 2004) it enabled structural and functional studies of significantly larger systems. The optimal field strengths for TROSY have been estimated to be on spectrometers operating around 900 MHz (21.14 T) for the 1HN TROSY (Pervushin et al. in Proc Natl Acad Sci USA 94:12366-12371, 1997) while the aromatic 13C (13Caro) TROSY is posited to be optimal at around 600 MHz (14.09 T) (Pervushin et al. in J Am Chem Soc 120:6394-6400, 1998b; Pervushin in Q Rev Biophys 33:161-197, 2000). The initial rational was based on the consideration of where the quadratic B0 field dependences of the TROSY relaxation rates reach a minimum. For sensitivity consideration, however, it is interesting to estimate which field strengths yield the tallest peaks. Recent studies of 15N-detected TROSYs suggested that maximal peak heights are expected at 1.15 GHz (27.01 T) although the slowest relaxation rates or longest transverse relaxation times T2 are indeed expected around 900 MHz (21.14 T) (Takeuchi in J Biomol NMR 63:323-331, 2015; Takeuchi et al. in J Biomol NMR 64:143-151, 2016). This was based on the fact that the heights of Lorentzian lines are proportional to B o3/2 * T2 (Bo). Thus, multiplying the parabolic T2(Bo) dependence with the increasing function of B o3/2 shifts the maxima of peak-height field dependence from the T2 maximum at 900 MHz to higher fields. Moreover, besides shifting the peak height maximum for 15N TROSY, this analysis yields estimates for optimal peak heights for 1HN detected TROSY to 1.5 GHz, and to 900 MHz for 13C-detected 13CaroTROSY as is detailed below. To our knowledge, this aspect of field dependence of TROSY sensitivity has not been in the attention of the NMR community but may affect perspectives of NMR at ultra-high fields.

Dataset Information

Nitrogen-detected TROSY yields comparable sensitivity to proton-detected TROSY for non-deuterated, large proteins under physiological salt conditions.

Publications

Nitrogen-detected TROSY yields comparable sensitivity to proton-detected TROSY for non-deuterated, large proteins under physiological salt conditions.

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