Transient dimer in the refolding kinetics of cytochrome c characterized by small-angle X-ray scattering.


Abstract

The equilibrium unfolding and the kinetic refolding of cytochrome c (Cyt c) in the presence of imidazole were studied with small-angle X-ray scattering (SAXS). The equilibrium unfolding experiments showed the radius of gyration, R(g), of native Cyt c to swell approximately 1 A with the addition of imidazole. The thermodynamic parameter m also reflects an expansion of the protein as its lower value demonstrates an increase in solvent-accessible surface area over that of native Cyt c in the absence of imidazole. Refolding was studied in the presence of imidazole as it prevents misligated intermediate states from forming during the refolding process, simplifying the kinetics, and making them easier to resolve. Time-resolved decreases in the forward scattering amplitude, I(0), demonstrated the transient formation of an aggregated intermediate. Final protein and denaturant concentrations were varied in the refolding kinetics, and the singular value decomposition (SVD) method was employed to characterize the associated state. This state was determined to be a dimer, with properties consistent with a molten globule. Study holds ProTherm entries: 23841, 23842 Extra Details: deltaGU = deltaGU - m[GdnHCl] where deltaGU is the change in free energy of state U relative to the native state at neutral pH and in the absence of denaturants and m is the linear dependence of deltaGU upon the denaturant concentration. Without Imidazole, SAXS= small-angle X-ray scattering; unfolding/refolding kinetics, small-angle x-ray scattering, cytochome c, singular value decomposition.

Submission Details

ID: esYcwWUT3

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:54 p.m.

Version: 1

Publication Details
Segel DJ;Eliezer D;Uversky V;Fink AL;Hodgson KO;Doniach S,Biochemistry (1999) Transient dimer in the refolding kinetics of cytochrome c characterized by small-angle X-ray scattering. PMID:10563821
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
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
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
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
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
5DF5 2016-09-14 1.3 The structure of oxidized rat cytochrome c (T28E) at 1.30 angstroms resolution.
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.
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
3O20 2012-01-25 1.9 Electron transfer complexes:experimental mapping of the Redox-dependent Cytochrome C electrostatic surface
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
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
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