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Journal of Biomolecular NMR

, Volume 3, Issue 6, pp 627–638 | Cite as

Measurement of amide proton exchange rates and NOEs with water in 13C/15N-enriched calcineurin B

  • Stephan Grzesiek
  • Ad Bax
Research Papers

Summary

A rapid and sensitive 2D approach is presented for measuring amide proton exchange rates and the NOE interaction between amide protons and water. The approach is applicable to uniformly 13C/15N-enriched proteins and can measure magnetization exchange rates in the 0.02 to >20s−1 range. The experiments rely on selective excitation of the water resonance, coupled with purging of underlying Hα resonances, followed by NOESY-or ROESY-type transfer to amide protons, which are dispersed by the amide 15N frequencies in an HSQC-type experiment. Two separate but interleaved experiments, with and without selective inversion of the H2O resonance, yield quantitative results. The method is demonstrated for a sample of the calcium-binding protein calcineurin B. Results indicate rapid amide exchange for the five calcineurin B residues that are analogous to the five rapidly exchanging residues in the ‘central helix’ of the homologous protein calmodulin.

Keywords

Hydrogen exchange NOESY ROESY Water Protein 2D NMR Calcineurin B 

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Copyright information

© ESCOM Science Publishers B.V 1993

Authors and Affiliations

  • Stephan Grzesiek
    • 1
  • Ad Bax
    • 1
  1. 1.Laboratory of Chemical Physics, National Institute of Diabetes and Digestive and Kidney DiseasesNational Institutes of HealthBethesdaUSA

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