Relationship between local and global stabilities of proteins: site-directed mutants and chemically-modified derivatives of cytochrome c.


Abstract

The methionine 80 sulfur-heme iron bond of rat cytochrome c, whose stability is decreased by mutating the phylogenetically invariant residue proline 30 to alanine and increased when tyrosine 67 is changed to phenylalanine, recovers its wild-type characteristics when both substitutions are performed on the same molecule. Titrations with urea, analyzed according to the heteropolymer theory [Alonso, D. O. V., & Dill, K. A. (1991) Biochemistry 30, 5974-5985], indicate that both single mutations increase the solvent exposure of hydrophobic groups in the unfolded state, while in the double mutant this conformational perturbation disappears. Similar increases in solvent exposure of hydrophobic groups are observed when the sulfur-iron bond of the wild-type protein is broken by alkylation of the methionine sulfur, by high pH, or by binding the heme iron with cyanide. The compensatory effects of the two single mutations do not extend to the overall stability of the protein. The added loss of conformational stability due to the single mutations amounts to 7.3 kcal/mol out of the 9 kcal/mol representing the overall free energy of stabilization of the native conformation of the wild-type protein. The folded conformation of the doubly mutated protein is only 2 kcal/mol less stable than that of the wild type. These results indicate that the double mutant protein is able to retain the essential folding pattern of cytochrome c and the thermodynamic stability of the methionine sulfur-heme iron bond, in spite of structural differences that weaken the overall stability of the molecule. Study holds ProTherm entries: 5320, 5321, 5322, 5323, 5324, 5325, 5326, 5327 Extra Details:

Submission Details

ID: fBKjDfUi

Submitter: Connie Wang

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

Version: 1

Publication Details
Schejter A;Luntz TL;Koshy TI;Margoliash E,Biochemistry (1992) Relationship between local and global stabilities of proteins: site-directed mutants and chemically-modified derivatives of cytochrome c. PMID:1326327
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
1OCD 1997-06-16 CYTOCHROME C (OXIDIZED) FROM EQUUS CABALLUS, NMR, MINIMIZED AVERAGE STRUCTURE
2N3B 2015-10-28 Structure of oxidized horse heart cytochrome c encapsulated in reverse micelles
2N9J 2016-02-17 Solution structure of oxidized human cytochrome c
1AKK 1997-09-17 SOLUTION STRUCTURE OF OXIDIZED HORSE HEART CYTOCHROME C, NMR, MINIMIZED AVERAGE STRUCTURE
5ZKV 2019-05-22 Solution structure of molten globule state of L94G mutant of horse cytochrome-c
1FI9 2000-08-23 SOLUTION STRUCTURE OF THE IMIDAZOLE COMPLEX OF CYTOCHROME C
2FRC 1997-07-29 CYTOCHROME C (REDUCED) FROM EQUUS CABALLUS, NMR, MINIMIZED AVERAGE STRUCTURE
1J3S 2004-05-18 Solution Structure of Reduced Recombinant Human Cytochrome c
1FI7 2000-08-23 Solution structure of the imidazole complex of cytochrome C
1LC2 2003-06-03 Solution Structure Of Reduced Horse Heart Cytochrome c in 30% Acetonitrile Solution, NMR 30 Structures
2N3Y 2016-12-14 NMR structure of the Y48pCMF variant of human cytochrome c in its reduced state
2GIW 1998-12-09 SOLUTION STRUCTURE OF REDUCED HORSE HEART CYTOCHROME C, NMR, 40 STRUCTURES
1LC1 2003-06-03 Solution Structure Of Reduced Horse Heart Cytochrome c in 30% Acetonitrile Solution, NMR Minimized Average Structure
2N9I 2016-02-17 Solution structure of reduced human cytochrome c
1M60 2002-08-07 Solution Structure of Zinc-substituted cytochrome c
1I5T 2001-03-21 SOLUTION STRUCTURE OF CYANOFERRICYTOCHROME C
1GIW 1998-12-09 SOLUTION STRUCTURE OF REDUCED HORSE HEART CYTOCHROME C, NMR, MINIMIZED AVERAGE STRUCTURE
5C0Z 2016-09-21 1.12 The structure of oxidized rat cytochrome c at 1.13 angstroms resolution
5DFS 2016-03-02 1.15 Crystal structure of Spider Monkey Cytochrome C at 1.15 Angstrom
5TY3 2017-09-06 1.25 Crystal structure of K72A variant of Human Cytochrome c
5DF5 2016-09-14 1.3 The structure of oxidized rat cytochrome c (T28E) at 1.30 angstroms resolution.
3ZOO 2013-10-23 1.35 Structure of the Y46F mutant of human cytochrome c
5O10 2018-03-28 1.36 Y48H mutant of human cytochrome c
5C9M 2016-09-21 1.36 The structure of oxidized rat cytochrome c (T28A) at 1.362 angstroms resolution.
2B4Z 2005-10-11 1.5 Crystal structure of cytochrome C from bovine heart at 1.5 A resolution.
5EXQ 2016-11-30 1.6 Human cytochrome c Y48H
3ZCF 2013-10-23 1.65 Structure of recombinant human cytochrome c
6FF5 2018-03-21 1.74 X-ray structure of bovine heart cytochrome c at high ionic strength
3O1Y 2012-01-25 1.75 Electron transfer complexes: Experimental mapping of the redox-dependent cytochrome c electrostatic surface
3WUI 2014-07-16 1.8 Dimeric horse cytochrome c formed by refolding from molten globule state
1WEJ 1998-12-09 1.8 IGG1 FAB FRAGMENT (OF E8 ANTIBODY) COMPLEXED WITH HORSE CYTOCHROME C AT 1.8 A RESOLUTION
3WC8 2013-12-11 1.8 Dimeric horse cytochrome c obtained by refolding with desalting method
6DUJ 2019-06-26 1.82 Crystal structure of A51V variant of Human Cytochrome c
3O20 2012-01-25 1.9 Electron transfer complexes:experimental mapping of the Redox-dependent Cytochrome C electrostatic surface
3NWV 2011-03-09 1.9 Human cytochrome c G41S
1HRC 1994-11-01 1.9 HIGH-RESOLUTION THREE-DIMENSIONAL STRUCTURE OF HORSE HEART CYTOCHROME C
2YBB 2011-10-19 19.0 Fitted model for bovine mitochondrial supercomplex I1III2IV1 by single particle cryo-EM (EMD-1876)
5IY5 2017-01-11 2.0 Electron transfer complex of cytochrome c and cytochrome c oxidase at 2.0 angstrom resolution
1CRC 1996-03-08 2.08 CYTOCHROME C AT LOW IONIC STRENGTH
3NBT 2010-07-14 2.1 Crystal structure of trimeric cytochrome c from horse heart
4NFG 2014-09-24 2.11 K13R mutant of horse cytochrome c and yeast cytochrome c peroxidase complex
4RSZ 2015-01-14 2.19 The X-ray structure of the Primary Adduct formed in the Reaction between Cisplatin and Cytochrome c
3NBS 2010-07-14 2.2 Crystal structure of dimeric cytochrome c from horse heart
1KTD 2002-05-01 2.4 CRYSTAL STRUCTURE OF CLASS II MHC MOLECULE IEK BOUND TO PIGEON CYTOCHROME C PEPTIDE
1U75 2004-09-28 2.55 Electron Transfer Complex between Horse Heart Cytochrome c and Zinc-Porphyrin Substituted Cytochrome c Peroxidase
6ECJ 2019-08-14 2.7 Human cytochrome c G41T
2PCB 1993-07-15 2.8 CRYSTAL STRUCTURE OF A COMPLEX BETWEEN ELECTRON TRANSFER PARTNERS, CYTOCHROME C PEROXIDASE AND CYTOCHROME C
3JBT 2015-11-18 3.8 Atomic structure of the Apaf-1 apoptosome
5JUY 2016-10-19 4.1 Active human apoptosome with procaspase-9
5WVE 2017-02-08 4.4 Apaf-1-Caspase-9 holoenzyme
3J2T 2013-04-10 9.5 An improved model of the human apoptosome

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
95.2 Cytochrome c, somatic B4USV4 CYC_OTOGA
95.1 Cytochrome c, somatic P00021 CYC_COLLI
96.1 Cytochrome c, somatic P00020 CYC_ANAPL
93.3 Cytochrome c, somatic P81280 CYC_ALLMI
92.4 Cytochrome c, somatic Q52V10 CYC_SAISC
95.2 Cytochrome c, somatic P00012 CYC_MIRLE
99.0 Cytochrome c, somatic Q52V09 CYC_CEPBA
95.2 Cytochrome c, somatic P00013 CYC_MINSC
95.2 Cytochrome c, somatic P00014 CYC_MACGI
96.2 Cytochrome c, somatic P00011 CYC_CANLF
100.0 Cytochrome c, somatic P62898 CYC_RAT
98.1 Cytochrome c, somatic P00008 CYC_RABIT
100.0 Cytochrome c, somatic P62897 CYC_MOUSE
98.1 Cytochrome c, somatic P68098 CYC_LAMGU
98.1 Cytochrome c, somatic P68100 CYC_ESCRO
98.1 Cytochrome c, somatic P68099 CYC_CAMDR
96.2 Cytochrome c, somatic 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
91.3 Cytochrome c, somatic P21665 CYC_VARVA
91.4 Cytochrome c, somatic Q640U4 CYC1_XENTR
91.3 Cytochrome c, somatic Q6IQM2 CYC_DANRE
90.5 Cytochrome c, somatic Q7YR71 CYC_TRACR
90.4 Cytochrome c, somatic Q6DKE1 CYC2_XENLA
91.4 Cytochrome c, somatic P00024 CYC_LITCT
91.4 Cytochrome c, somatic P00022 CYC_CHESE
91.4 Cytochrome c, somatic Q5RFH4 CYC_PONAB
91.4 Cytochrome c, somatic P99998 CYC_PANTR
91.4 Cytochrome c, somatic P99999 CYC_HUMAN
91.4 Cytochrome c, somatic Q6WUX8 CYC_GORGO
90.5 Cytochrome c, somatic P00003 CYC_ATESP
92.4 Cytochrome c, somatic P00019 CYC_STRCA
92.4 Cytochrome c, somatic Q52V08 CYC_MACSY
92.4 Cytochrome c, somatic P00002 CYC_MACMU
92.4 Cytochrome c, somatic P00018 CYC_DRONO
92.4 Cytochrome c, somatic P67882 CYC_MELGA
92.4 Cytochrome c, somatic P67881 CYC_CHICK
92.4 Cytochrome c, somatic P00017 CYC_APTPA