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  Optimum levels of exchangeable protons in perdeuterated proteins for proton detection in MAS solid-state NMR spectroscopy.

Akbey, Ü., Lange, S., Franks, W. T., Linser, R., Rehbein, K., Diehl, A., et al. (2010). Optimum levels of exchangeable protons in perdeuterated proteins for proton detection in MAS solid-state NMR spectroscopy. Journal of Biomolecular NMR, 46(1), 67-73. doi:10.1007/s10858-009-9369-0.

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Item Permalink: http://hdl.handle.net/11858/00-001M-0000-0018-EB2B-1 Version Permalink: http://hdl.handle.net/11858/00-001M-0000-0028-DD53-0
Genre: Journal Article

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 Creators:
Akbey, Ü., Author
Lange, S., Author
Franks, W. T., Author
Linser, R.1, Author              
Rehbein, K., Author
Diehl, A., Author
van Rossum, B. J., Author
Reif, B., Author
Oschkinat, H., Author
Affiliations:
1Research Group of Solid-State NMR-2, MPI for Biophysical Chemistry, Max Planck Society, ou_1950286              

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Free keywords: Proton detection; Deuteration; Protein; MAS NMR; alpha-spectrin SH3
 Abstract: We present a systematic study of the effect of the level of exchangeable protons on the observed amide proton linewidth obtained in perdeuterated proteins. Decreasing the amount of D2O employed in the crystallization buffer from 90 to 0%, we observe a fourfold increase in linewidth for both 1H and 15N resonances. At the same time, we find a gradual increase in the signal-to-noise ratio (SNR) for 1H–15N correlations in dipolar coupling based experiments for H2O concentrations of up to 40%. Beyond 40%, a significant reduction in SNR is observed. Scalar-coupling based 1H–15N correlation experiments yield a nearly constant SNR for samples prepared with ≤30% H2O. Samples in which more H2O is employed for crystallization show a significantly reduced NMR intensity. Calculation of the SNR by taking into account the reduction in 1H T 1 in samples containing more protons (SNR per unit time), yields a maximum SNR for samples crystallized using 30 and 40% H2O for scalar and dipolar coupling based experiments, respectively. A sensitivity gain of 3.8 is obtained by increasing the H2O concentration from 10 to 40% in the CP based experiment, whereas the linewidth only becomes 1.5 times broader. In general, we find that CP is more favorable compared to INEPT based transfer when the number of possible 1H,1H interactions increases. At low levels of deuteration (≥60% H2O in the crystallization buffer), resonances from rigid residues are broadened beyond detection. All experiments are carried out at MAS frequency of 24 kHz employing perdeuterated samples of the chicken α-spectrin SH3 domain.

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Language(s): eng - English
 Dates: 2009-08-222010-01
 Publication Status: Published in print
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 Rev. Method: Peer
 Identifiers: DOI: 10.1007/s10858-009-9369-0
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Title: Journal of Biomolecular NMR
Source Genre: Journal
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Pages: - Volume / Issue: 46 (1) Sequence Number: - Start / End Page: 67 - 73 Identifier: -