Fractional enrichment of proteins using [2-13C]-glycerol as the carbon source facilitates measurement of excited state 13Cα chemical shifts with improved sensitivity

At the time for thesis presentation publication was in status: ManuscriptAt the time for thesis presentation name of publication was: Fractional enrichment using [2-13C]-glycerol as the carbon source facilitates measurements of excited state 13Cα chemical shifts with improved sensitivity
A selective isotope labeling scheme based on the utilization of [2-13C]-glycerol as the carbon source during protein overexpression has been evaluated for the measurement of excited state 13Cα 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 [2-13C]-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 13Cα 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 13Cα–13Cβ spin-pairs that are present for many residue types. The labeling approach described here should thus be attractive for studies of exchanging systems using 13Cα spin probes.

Andresen, CeciliaLinköpings universitet,Kemi,Tekniska högskolan (author)
Khan, Shahid N.Linköpings universitet,Kemi,Tekniska högskolan (author)
Kay, Lewis E.Departments of Medical Genetics, Biochemistry and Chemistry, The University of Toronto, Canada (author)
Lundström, PatrikLinköpings universitet,Kemi,Tekniska högskolan(Swepub:liu)patlu71 (author)
Linköpings universitetKemi (creator_code:org_t)

In:Journal of Biomolecular NMR: Springer Netherlands62:3, s. 341-3510925-27381573-5001

By the author/editor: Ahlner, Alexandr ...; Andresen, Cecili ...; Khan, Shahid N.; Kay, Lewis E.; Lundström, Patri ...

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