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Cited by 76Lipid Membrane Structure and BehaviorAdvanced NMR Techniques and ApplicationsProtein Structure and Dynamics

Cytochrome-c interactions with cardiolipin in bilayers – a multinuclear magic-angle spinning NMR-study

Spooner, Paul J. R., Watts, Anthony·1992·Biochemistry
DOI 10.1021/bi00156a037
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Abstract

The influence of cytochrome c binding to cardiolipin bilayers on the motional characteristics of each component has been analyzed by magic-angle spinning (MAS) NMR. Observations were made by NMR of natural abundance 31P, 13C, and 1H nuclei in the lipid as well as sites enriched with 13C in the protein. Analysis of methyl carbons enriched in ([epsilon-13CH3]methionine)cytochrome c at residues 65 and 80 reveal quite different behavior for these sites when the protein was bound at a 1:15 molar ratio with hydrated cardiolipin. Cross-polarization (CP) shows a single broad resonance downfield in the methyl region which corresponds to the spectral characteristics of methionine 65 in the solution protein when subjected to moderate thermal perturbations. These observations suggest that although methionine 65 remains motionally restricted when the protein binds to the lipid bilayers, this residue becomes less shielded and exposed to more chemically distinct environments than in the native state of the protein. In contrast to its behavior in native oxidized protein, the methionine 80 methyl could be detected following direct pi/2 pulse excitation, and this residue is assumed to be released from the axial ligand site on the heme iron to become more exposed and highly mobile in the protein-lipid complex. An analysis of the CP response for natural abundance 13C nuclei in the lipid reveals a general increase in motions with slower rates (tens of kilohertz) on binding with cytochrome c, except for sites within the region of fatty acyl chain unsaturation which appear to be selectively mobilized in the complex with protein. It is concluded that, aside from effects on the unsaturated segments, the bound protein induces new modes of slow motions in the lipid assemblies rather than restricting the overall reorientation freedom of the lipid. The strong paramagnetic effects observed previously on the relaxation of phosphorus in protein-bound lipid [Spooner, P.J.R., & Watts, A. (1991) Biochemistry 30, 3880-3885] were not extended to any carbon and proton sites observable by MAS NMR in the lipid, and this infers a specific interaction of lipid phosphate groups with the heme. However, when protein was bound to cardiolipin mixed at a 1:4 mole ratio with dioleoylphosphatidylcholine in bilayers, no direct interaction with the heme was apparent from the phosphorus NMR relaxation behavior in this component, resolved by MAS. Instead, the spectral anisotropy of cardiolipin phosphorus was determined to be reduced, indicating that, on binding with cytochrome c, the headgroup organization was perturbed in this component.(ABSTRACT TRUNCATED AT 400 WORDS)

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  • The heme iron of cytochrome c is predominately hexacoordinate, but can become pentacoordinate when the iron-methionine bond is ruptured which occurs when the methionine is oxidized, tyrosines are nitrated, or the cytochrome interacts with anionic phospholipids.

    “Similar to Ngb, the heme iron of cytochrome c is predominately hexacoordinate, but can become pentacoordinate when the iron-methionine bond is ruptured which occurs when the methionine is oxidized [65, 66], tyrosines are nitrated [67, 68], or the cytochrome interacts with anionic phospholipids such as those present in the inner membrane of the mitochondria [69–71].”

    From Mechanisms of nitrite bioactivation (2014) · cites this paper as [69]

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Authors
Spooner, P. J. R., & Watts, A.
Affiliations
University of Oxford (GB)
ORCID
Watts, Anthony
Venue
Biochemistry
Publisher
American Chemical Society (ACS)
Published
1992-10-01
Volume
31
Issue
41
Pages
10129-10138
ISSN
  • 0006-2960 (ISSN-L)
  • 1520-4995
DOI
10.1021/bi00156a037
OpenAlex
W2058982127
Cited by
76
References
45
Citations by year
2025: 2 · 2024: 1 · 2023: 1 · 2020: 2 · 2019: 2 · 2018: 3 · 2017: 3 · 2016: 1 · 2015: 2 · 2014: 2 · 2013: 4 · 2012: 1
FWCI
1.98
Type
journal article
Language
en
Open access
closed
Locations
  • Biochemistry (publishedVersion)
  • PubMed (publishedVersion)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Lipid Membrane Structure and Behavior
  • Physical Sciences › Chemistry › Spectroscopy › Advanced NMR Techniques and Applications
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein Structure and Dynamics
Keywords
cytochrome c, cardiolipin, protein-lipid interactions, lipid bilayers, heme axial ligand, lipid mobility, paramagnetic effects, DOPC
MeSH terms
Carbon Isotopes, Cardiolipins, Cytochrome c Group, Lipid Bilayers, Magnetic Resonance Spectroscopy

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