Overexpression of myoglobin and assignment of its amide, Cα and Cβ resonances

Patricia A. Jennings, Martin J. Stone, Peter E. Wright

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Abstract

Sperm whale apomyoglobin was expressed to high levels on minimal media and isotopically labeled with 13C and 15N nuclei. The isotopically labeled apoprotein was purified to homogeneity in a single step by reversed-phase chromatography and reconstituted with hemin and carbon monoxide gas for NMR analysis. Sequence-specific backbone 1HN, 15N and 13Cα as well as side-chain 13Cβ resonance assignments have been made for over 90% of the amino acids in the carbon monoxide complex of the protein. Resonance assignments were made by analysis of a series of 3D triple resonance spectra measured on the uniformly labeled sample. These assignments will provide the basis for analyzing the effects of point site mutations on the structure, stability and dynamics of the protein in solution.

Original languageEnglish
Pages (from-to)271-276
Number of pages6
JournalJournal of Biomolecular NMR
Volume6
Issue number3
DOIs
Publication statusPublished - Nov 1995
Externally publishedYes

Keywords

  • Heme protein
  • Heteronuclear NMR
  • Inclusion bodies
  • Myoglobin

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