Backbone and side-chain heteronuclear resonance assignments and hyperfine NMR shifts in horse cytochrome c

Weixia Liu, Jon Rumbley, S. Walter Englander, A. Joshua Wand

Research output: Contribution to journalArticlepeer-review

25 Scopus citations

Abstract

The [H26N, H33N] mutant of horse heart cytochrome c was expressed in E. coli during growth on isotopically enriched minimal media. Complete resonance assignments of both the diamagnetic reduced (spin zero) and paramagnetic oxidized (spin 1/2) states of the protein were obtained using standard triple resonance and total correlation spectroscopy using the previously determined 1H chemical shifts of the wild-type protein as a guide. The correspondence of chemical shifts between the wild type and the mutant protein is excellent, indicating that they have nearly identical structures. The expanded library of chemical shifts for both redox states in both proteins allowed the refinement of the electron spin g-tensor of the oxidized states. The g-tensors of the oxidized states of the wild-type and [H26N, H33N] mutant proteins are closely similar, indicating that the subtle details of the ligand fields are nearly identical. The refined g-tensors were then used to probe for redox-dependent structure change in the two proteins.

Original languageEnglish (US)
Pages (from-to)2104-2108
Number of pages5
JournalProtein Science
Volume12
Issue number9
DOIs
StatePublished - Sep 1 2003

Keywords

  • G-tensor
  • Hyperfine shifts
  • Labeling hemeproteins
  • NMR resonance assignments
  • Paramagnetic shifts

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