Tertiary stability of native and methionine-80 modified cytochrome c detected by proton-deuterium exchange using on-line Fourier transform infrared spectroscopy.


The stability of the tertiary structure of cytochrome c and of a methionine-80 chemically modified form of this protein has been investigated by monitoring on-line the exchange of amide protons with deuterons using infrared spectroscopy. The modified protein has a structural stabilization energy of approximately 50% of that of native cytochrome c, whereas the secondary structure is not affected by the modification. In the modified protein the fraction of slow exchanging amides is smaller compared to that in the native protein, and the exchange rate constants are found to be 2-3 times larger for the slow (half-life of 5.5 h) and intermediate (half-life of 4.1 min) exchanging fraction of amides. The exchange rate of a fast exchanging fraction of amides (half-life smaller than 1 min), most likely surface exposed amides, is not influenced by tertiary destabilization of the protein. The results in aqueous solution agree well with data obtained by monitoring the amide-proton exchange using 1H-nuclear magnetic resonance. In films, using attenuated total reflection infrared techniques, this difference in tertiary stability between modified and native cytochrome c could also be demonstrated. The various advantages and complications of this approach are discussed in detail. Study holds ProTherm entries: 5293, 5294, 5295 Extra Details: secondary structure; exchange rate constants; surface exposed;,tertiary stability

Submission Details

ID: ee58GhrJ3

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:29 p.m.

Version: 1

Publication Details
de Jongh HH;Goormaghtigh E;Ruysschaert JM,Biochemistry (1995) Tertiary stability of native and methionine-80 modified cytochrome c detected by proton-deuterium exchange using on-line Fourier transform infrared spectroscopy. PMID:7819193
Additional Information

Structure view and single mutant data analysis

Study data

No weblogo for data of varying length.
Colors: D E R H K S T N Q A V I L M F Y W C G P

Data Distribution

Studies with similar sequences (approximate matches)

Correlation with other assays (exact sequence matches)

Relevant PDB Entries

Structure ID Release Date Resolution Structure Title
2B4Z 2005-09-27T00:00:00+0000 1.5 Crystal structure of cytochrome C from bovine heart at 1.5 A resolution.
2YBB 2011-03-02T00:00:00+0000 19.0 Fitted model for bovine mitochondrial supercomplex I1III2IV1 by single particle cryo-EM (EMD-1876)
3J2T 2012-12-23T00:00:00+0000 9.5 An improved model of the human apoptosome
5C0Z 2015-06-12T00:00:00+0000 1.12 The structure of oxidized rat cytochrome c at 1.13 angstroms resolution
5C9M 2015-06-28T00:00:00+0000 1.36 The structure of oxidized rat cytochrome c (T28A) at 1.362 angstroms resolution.
5DF5 2015-08-26T00:00:00+0000 1.3 The structure of oxidized rat cytochrome c (T28E) at 1.30 angstroms resolution.
5JUY 2016-05-10T00:00:00+0000 4.1 Active human apoptosome with procaspase-9
6FF5 2018-01-03T00:00:00+0000 1.74 X-ray structure of bovine heart cytochrome c at high ionic strength
6N1O 2018-11-09T00:00:00+0000 1.55 Oxidized rat cytochrome c mutant (S47E)

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
91.3 Cytochrome c P00021 CYC_COLLI
92.2 Cytochrome c P00020 CYC_ANAPL
92.3 Cytochrome c P81280 CYC_ALLMI
90.5 Cytochrome c Q52V10 CYC_SAISC
93.3 Cytochrome c P00012 CYC_MIRLE
93.3 Cytochrome c Q52V09 CYC_CEPBA
93.3 Cytochrome c P00013 CYC_MINSC
93.3 Cytochrome c P00014 CYC_MACGI
94.3 Cytochrome c P00011 CYC_CANLF
94.3 Cytochrome c P62898 CYC_RAT
94.3 Cytochrome c P00008 CYC_RABIT
94.3 Cytochrome c P62897 CYC_MOUSE
95.2 Cytochrome c P68098 CYC_LAMGU
95.2 Cytochrome c P68100 CYC_ESCRO
95.2 Cytochrome c P68099 CYC_CAMDR
94.3 Cytochrome c P00007 CYC_HIPAM
97.1 Cytochrome c P62896 CYC_SHEEP
97.1 Cytochrome c P62895 CYC_PIG
97.1 Cytochrome c P62894 CYC_BOVIN
99.0 Cytochrome c P68096 CYC_EQUBU
99.0 Cytochrome c P68097 CYC_EQUAS
100.0 Cytochrome c P00004 CYC_HORSE
90.3 Cytochrome c B4USV4 CYC_OTOGA