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The response of Ω-loop D dynamics to truncation of trimethyllysine 72 of yeast iso-1-cytochrome c depends on the nature of loop deformation

Abstract

Trimethyllysine 72 (tmK72) has been suggested to play a role in sterically constraining the heme crevice dynamics of yeast iso-1-cytochrome c mediated by the Ω-loop D cooperative substructure (residues 70–85). A tmK72A mutation causes a gain in peroxidase activity, a function of cytochrome c that is important early in apoptosis. More than one higher energy state is accessible for the Ω-loop D substructure via tier 0 dynamics. Two of these are alkaline conformers mediated by Lys73 and Lys79. In the current work, the effect of the tmK72A mutation on the thermodynamic and kinetic properties of wild-type iso-1-cytochrome c (yWT versus WT*) and on variants carrying a K73H mutation (yWT/K73H versus WT*/K73H) is studied. Whereas the tmK72A mutation confers increased peroxidase activity in wild-type yeast iso-1-cytochrome c and increased dynamics for formation of a previously studied His79-heme alkaline conformer, the tmK72A mutation speeds return of the His73-heme alkaline conformer to the native state through destabilization of the His73-heme alkaline conformer relative to the native conformer. These opposing behaviors demonstrate that the response of the dynamics of a protein substructure to mutation depends on the nature of the perturbation to the substructure. For a protein substructure which mediates more than one function of a protein through multiple non-native structures, a mutation could change the partitioning between these functions. The current results suggest that the tier 0 dynamics of Ω-loop D that mediates peroxidase activity has similarities to the tier 0 dynamics required to form the His79-heme alkaline conformer.

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Abbreviations

ET:

Electron transfer

gated ET:

Conformationally gated electron transfer

GdnHCl:

Guanidine hydrochloride

iso-1-Cytc :

Iso-1-cytochrome c

tmK:

Trimethyllysine

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Acknowledgments

This research was supported by NSF Grants CHE-0910166 and CHE-1306903 to BEB. The Bruker microflex MALDI-ToF mass spectrometer was acquired with support from an NSF Major Research Instrumentation Grant CHE-1039814. BEB also acknowledges support from CoBRE Grant P20GM103546 funded by the NIGMS.

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Correspondence to Bruce E. Bowler.

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McClelland, L.J., Seagraves, S.M., Khan, M.K.A. et al. The response of Ω-loop D dynamics to truncation of trimethyllysine 72 of yeast iso-1-cytochrome c depends on the nature of loop deformation. J Biol Inorg Chem 20, 805–819 (2015). https://doi.org/10.1007/s00775-015-1267-1

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Keywords

  • Cooperative substructure dynamics
  • Cytochrome c
  • Alkaline conformational transition
  • Conformationally gated electron transfer
  • Apoptosis