Title |
Fractional enrichment of proteins using [2-13C]-glycerol as the carbon source facilitates measurement of excited state 13Cα chemical shifts with improved sensitivity
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Published in |
Journal of Biomolecular NMR, May 2015
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DOI | 10.1007/s10858-015-9948-1 |
Pubmed ID | |
Authors |
Alexandra Ahlner, Cecilia Andresen, Shahid N. Khan, Lewis E. Kay, Patrik Lundström |
Abstract |
A selective isotope labeling scheme based on the utilization of [2-(13)C]-glycerol as the carbon source during protein overexpression has been evaluated for the measurement of excited state (13)Cα chemical shifts using Carr-Purcell-Meiboom-Gill (CPMG) relaxation dispersion (RD) experiments. As expected, the fractional incorporation of label at the Cα positions is increased two-fold relative to labeling schemes based on [2-(13)C]-glucose, effectively doubling the sensitivity of NMR experiments. Applications to a binding reaction involving an SH3 domain from the protein Abp1p and a peptide from the protein Ark1p establish that accurate excited state (13)Cα chemical shifts can be obtained from RD experiments, with errors on the order of 0.06 ppm for exchange rates ranging from 100 to 1000 s(-1), despite the small fraction of (13)Cα-(13)Cβ spin-pairs that are present for many residue types. The labeling approach described here should thus be attractive for studies of exchanging systems using (13)Cα spin probes. |
Mendeley readers
Geographical breakdown
Country | Count | As % |
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Unknown | 17 | 100% |
Demographic breakdown
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Student > Ph. D. Student | 4 | 24% |
Other | 2 | 12% |
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Chemical Engineering | 1 | 6% |
Other | 0 | 0% |
Unknown | 1 | 6% |