Unfolding-refolding behaviour of chicken egg white ovomucoid and its correlation with the three domain structure of the protein.


Abstract

The urea and heat-induced unfolding-refolding behaviours of chicken egg white ovomucoid and its four fragments representing domains I, II + III, I + II and III were systematically investigated in 0.06 M sodium phosphate buffer (pH 7.0) by difference spectral measurements. The effect of temperature on ovomucoid and its fragments was also studied in 0.05 M sodium acetate buffer (pH 5.0) and in presence of 2 M urea at pH 7.0. Intrinsic viscosity data showed that ovomucoid and its different fragments did not lose any significant amount of their structure under mild acidic conditions (pH 4.6). Difference spectral results showed extensive disruption of the native structure by urea or temperature. Isothermal transitions showed single-step for domain I, domain I + II and domain III, and two-step having one stable intermediate, for ovomucoid and its fragment representing domain II + III. However, the presence of intermediate was not detected when the transitions were studied with temperature at pH 7.0. Strikingly, the single-step thermal transitions of ovomucoid and its fragment representing domain II + III, became two-step when measured either at pH 5.0 or in presence of 2 M urea at pH 7.0. Analysis of the equilibrium data on urea and heat denaturation showed that the second transition observed with ovomucoid or domain II + III represent the unfolding of domain III. The kinetic results of ovomucoid and its fragments indicate that the protein unfolds with three kinetic phases. A comparison of three rate constants for the unfolding of intact ovomucoid with that of its various fragments revealed that domain I, II and III of the protein correspond to the three kinetic phases having rate constants 0.456, 0.120 and 0.054 min-1, respectively. These data have led us to conclude: (i) the unusual stability of ovomucoid towards various denaturants, including temperature, is due to its domain III, (ii) initiation of the folding of the ovomucoid molecule starts from its NH2-terminal region which probably provides the nucleation site for the formation of the subsequent structure and (iii) domains I and II have greater mutual recognition between them as compared to the recognition either of them have with domain III. Study holds ProTherm entries: 10066, 10067, 10068, 10069, 10070, 10071, 10072, 10073, 10074, 10075, 10076, 10077, 10078, 10080, 10081, 10082, 10083, 10084 Extra Details: transitionI ovomucoid; egg-white trypsin inhibitor; structual domain;,conformation; equilibrium studies; kinetic studies; protein denaturation

Submission Details

ID: YxnzuNjU

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:39 p.m.

Version: 1

Publication Details
Das BK;Agarwal SK;Khan MY,Biochim. Biophys. Acta (1991) Unfolding-refolding behaviour of chicken egg white ovomucoid and its correlation with the three domain structure of the protein. PMID:2001382
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
1M8B 2002-09-04 Solution structure of the C State of turkey ovomucoid at pH 2.5
1M8C 2002-09-04 SOLUTION STRUCTURE OF THE T State OF TURKEY OVOMUCOID AT PH 2.5
1TUR 1994-10-15 SOLUTION STRUCTURE OF TURKEY OVOMUCOID THIRD DOMAIN AS DETERMINED FROM NUCLEAR MAGNETIC RESONANCE DATA
1IY6 2003-03-11 Solution structure of OMSVP3 variant, P14C/N39C
1IY5 2003-03-11 Solution structure of wild type OMSVP3
1TUS 1994-10-15 SOLUTION STRUCTURE OF REACTIVE-SITE HYDROLYZED TURKEY OVOMUCOID THIRD DOMAIN BY NUCLEAR MAGNETIC RESONANCE AND DISTANCE GEOMETRY METHODS
1OMU 1996-03-08 SOLUTION STRUCTURE OF OVOMUCOID (THIRD DOMAIN) FROM DOMESTIC TURKEY (298K, PH 4.1) (NMR, 50 STRUCTURES) (REFINED MODEL USING NETWORK EDITING ANALYSIS)
1OMT 1996-03-08 SOLUTION STRUCTURE OF OVOMUCOID (THIRD DOMAIN) FROM DOMESTIC TURKEY (298K, PH 4.1) (NMR, 50 STRUCTURES) (STANDARD NOESY ANALYSIS)
1R0R 2003-11-11 1.1 1.1 Angstrom Resolution Structure of the Complex Between the Protein Inhibitor, OMTKY3, and the Serine Protease, Subtilisin Carlsberg
2GKR 2007-02-13 1.16 Crystal structure of the N-terminally truncated OMTKY3-del(1-5)
2GKT 2007-02-13 1.23 Crystal structure of the P14'-Ala32 variant of the N-terminally truncated OMTKY3-del(1-5)
1SGP 1995-10-15 1.4 ALA 18 VARIANT OF TURKEY OVOMUCOID INHIBITOR THIRD DOMAIN COMPLEXED WITH STREPTOMYCES GRISEUS PROTEINASE B
2OVO 1985-11-08 1.5 THE CRYSTAL AND MOLECULAR STRUCTURE OF THE THIRD DOMAIN OF SILVER PHEASANT OVOMUCOID (OMSVP3)
1YU6 2005-05-03 1.55 Crystal Structure of the Subtilisin Carlsberg:OMTKY3 Complex
1CT4 2000-01-12 1.6 CRYSTAL STRUCTURE OF THE OMTKY3 P1 VARIANT OMTKY3-VAL18I IN COMPLEX WITH SGPB
2NU2 2006-11-21 1.65 Accommodation of positively-charged residues in a hydrophobic specificity pocket: Crystal structures of SGPB in complex with OMTKY3 variants Lys18I and Arg18I
1CT2 2000-01-12 1.65 CRYSTAL STRUCTURE OF THE OMTKY3 P1 VARIANT OMTKY3-THR18I IN COMPLEX WITH SGPB
1DS3 2001-01-31 1.65 CRYSTAL STRUCTURE OF OMTKY3-CH2-ASP19I
2GKV 2007-02-13 1.7 Crystal structure of the SGPB:P14'-Ala32 OMTKY3-del(1-5) complex
1DS2 2001-01-31 1.7 CRYSTAL STRUCTURE OF SGPB:OMTKY3-COO-LEU18I
2SGF 2003-08-26 1.75 PHE 18 VARIANT OF TURKEY OVOMUCOID INHIBITOR THIRD DOMAIN COMPLEXED WITH STREPTOMYCES GRISEUS PROTEINASE B
2NU4 2006-11-21 1.75 Accommodation of positively-charged residues in a hydrophobic specificity pocket: Crystal structures of SGPB in complex with OMTKY3 variants Lys18I and Arg18I
2SGP 2001-01-31 1.8 PRO 18 VARIANT OF TURKEY OVOMUCOID INHIBITOR THIRD DOMAIN COMPLEXED WITH STREPTOMYCES GRISEUS PROTEINASE B AT PH 6.5
1CHO 1988-07-16 1.8 CRYSTAL AND MOLECULAR STRUCTURES OF THE COMPLEX OF ALPHA-*CHYMOTRYPSIN WITH ITS INHIBITOR TURKEY OVOMUCOID THIRD DOMAIN AT 1.8 ANGSTROMS RESOLUTION
2NU3 2006-11-21 1.8 Accommodation of positively-charged residues in a hydrophobic specificity pocket: Crystal structures of SGPB in complex with OMTKY3 variants Lys18I and Arg18I
1SGY 2003-08-26 1.8 TYR 18 VARIANT OF TURKEY OVOMUCOID INHIBITOR THIRD DOMAIN COMPLEXED WITH STREPTOMYCES GRISEUS PROTEINASE B AT PH 6.5
1SGE 2003-08-26 1.8 GLU 18 VARIANT OF TURKEY OVOMUCOID INHIBITOR THIRD DOMAIN COMPLEXED WITH STREPTOMYCES GRISEUS PROTEINASE B AT PH 6.5
1SGN 2003-08-26 1.8 ASN 18 VARIANT OF TURKEY OVOMUCOID INHIBITOR THIRD DOMAIN COMPLEXED WITH STREPTOMYCES GRISEUS PROTEINASE B
1SGR 1995-10-15 1.8 LEU 18 VARIANT OF TURKEY OVOMUCOID INHIBITOR THIRD DOMAIN COMPLEXED WITH STREPTOMYCES GRISEUS PROTEINASE B
2NU1 2006-11-21 1.8 Molecular structures of the complexes of SGPB with OMTKY3 aromatic P1 variants Trp18I, His18I, Phe18I and Tyr18I
2SGQ 2003-08-26 1.8 GLN 18 VARIANT OF TURKEY OVOMUCOID INHIBITOR THIRD DOMAIN COMPLEXED WITH STREPTOMYCES GRISEUS PROTEINASE B AT PH 6.5
3SGB 1983-07-12 1.8 STRUCTURE OF THE COMPLEX OF STREPTOMYCES GRISEUS PROTEASE B AND THE THIRD DOMAIN OF THE TURKEY OVOMUCOID INHIBITOR AT 1.8 ANGSTROMS RESOLUTION
1SGD 2003-08-26 1.8 ASP 18 VARIANT OF TURKEY OVOMUCOID INHIBITOR THIRD DOMAIN COMPLEXED WITH STREPTOMYCES GRISEUS PROTEINASE B AT PH 6.5
2SGE 2003-08-26 1.8 GLU 18 VARIANT OF TURKEY OVOMUCOID INHIBITOR THIRD DOMAIN COMPLEXED WITH STREPTOMYCES GRISEUS PROTEINASE B AT PH 10.7
3SGQ 2003-08-26 1.8 GLN 18 VARIANT OF TURKEY OVOMUCOID INHIBITOR THIRD DOMAIN COMPLEXED WITH STREPTOMYCES GRISEUS PROTEINASE B AT PH 10.7
1CT0 2000-01-12 1.8 CRYSTAL STRUCTURE OF THE OMTKY3 P1 VARIANT OMTKY3-SER18I IN COMPLEX WITH SGPB
1PPF 1994-01-31 1.8 X-RAY CRYSTAL STRUCTURE OF THE COMPLEX OF HUMAN LEUKOCYTE ELASTASE (PMN ELASTASE) AND THE THIRD DOMAIN OF THE TURKEY OVOMUCOID INHIBITOR
2SGD 2003-08-26 1.8 ASP 18 VARIANT OF TURKEY OVOMUCOID INHIBITOR THIRD DOMAIN COMPLEXED WITH STREPTOMYCES GRISEUS PROTEINASE B AT PH 10.7
1Z7K 2005-04-05 1.9 Crystal Structure of Trypsin- Ovomucoid turkey egg white inhibitor complex
1CSO 2000-01-12 1.9 CRYSTAL STRUCTURE OF THE OMTKY3 P1 VARIANT OMTKY3-ILE18I IN COMPLEX WITH SGPB
1SGQ 1995-10-15 1.9 GLY 18 VARIANT OF TURKEY OVOMUCOID INHIBITOR THIRD DOMAIN COMPLEXED WITH STREPTOMYCES GRISEUS PROTEINASE B
2NU0 2006-11-21 1.95 Molecular structures of the complexes of SGPB with OMTKY3 aromatic P1 variants Trp18I, His18I, Phe18I, and Tyr18I
1HJA 1998-01-14 2.3 LYS 18 VARIANT OF TURKEY OVOMUCOID INHIBITOR THIRD DOMAIN COMPLEXED WITH ALPHA-CHYMOTRYPSIN
4OVO 1993-01-15 2.5 REFINED X-RAY CRYSTAL STRUCTURES OF THE REACTIVE SITE MODIFIED OVOMUCOID INHIBITOR THIRD DOMAINS FROM SILVER PHEASANT (OMSVP3(ASTERISK)) AND FROM JAPANESE QUAIL (OMJPQ3(ASTERISK))

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
90.4 Ovomucoid P05567 IOVO_ANSSE
90.4 Ovomucoid P52260 IOVO_GUTPU
90.4 Ovomucoid P05571 IOVO_COSCO
90.4 Ovomucoid P05570 IOVO_DENVD
90.4 Ovomucoid P68393 IOVO_BRACA
90.4 Ovomucoid P68394 IOVO_ANSCA
90.6 Ovomucoid P05595 IOVO_PELCO
90.4 Ovomucoid P68132 IOVO_OXYVI
90.4 Ovomucoid P68131 IOVO_OXYJA
90.4 Ovomucoid P68130 IOVO_CYGOL
90.4 Ovomucoid P68129 IOVO_CYGAT
92.3 Ovomucoid P67892 IOVO_DENBI
92.3 Ovomucoid P67891 IOVO_DENAU
92.3 Ovomucoid P67889 IOVO_DENAR
92.3 Ovomucoid P67890 IOVO_DENAC
92.5 Ovomucoid P05594 IOVO_PTEAF
94.2 Ovomucoid P68468 IOVO_NUMME
94.2 Ovomucoid P68469 IOVO_ACRVU
98.2 Ovomucoid P52267 IOVO_GALVA
92.5 Ovomucoid P05601 IOVO_ARBTO
94.6 Ovomucoid P05598 IOVO_FRAPO
94.5 Ovomucoid P05591 IOVO_COLVI
96.2 Ovomucoid P68147 IOVO_ALERU
96.2 Ovomucoid P68146 IOVO_ALECH
92.7 Ovomucoid P05596 IOVO_PTEER
100.0 Ovomucoid P01005 IOVO_CHICK
94.5 Ovomucoid P05588 IOVO_CALSP
96.3 Ovomucoid P52269 IOVO_SYRMI
92.9 Ovomucoid P68471 IOVO_PHAVE
92.9 Ovomucoid P68470 IOVO_PHACO
96.4 Ovomucoid P05589 IOVO_CALSC
96.4 Ovomucoid P05587 IOVO_OREPI
94.6 Ovomucoid P05605 IOVO_SYRRE
96.4 Ovomucoid P05592 IOVO_CYRMO
98.1 Ovomucoid P52263 IOVO_PAVMU
94.6 Ovomucoid P67894 IOVO_LOPIG
94.6 Ovomucoid P67893 IOVO_LOPDI
94.6 Ovomucoid P05585 IOVO_BONUM
94.6 Ovomucoid P05599 IOVO_PERPE
98.2 Ovomucoid P68388 IOVO_CALGM
98.2 Ovomucoid P68387 IOVO_CALCC
96.4 Ovomucoid P52258 IOVO_AFRCO
96.4 Ovomucoid P52262 IOVO_LOPLE
96.4 Ovomucoid P52261 IOVO_LOPBU
98.2 Ovomucoid P05609 IOVO_PAVCR
96.4 Ovomucoid P68127 IOVO_CHRPC
96.4 Ovomucoid P68128 IOVO_CHRAM
98.2 Ovomucoid P67944 IOVO_TYMCU
98.2 Ovomucoid P67949 IOVO_TRATE
98.2 Ovomucoid P67948 IOVO_TRASA
98.2 Ovomucoid P67947 IOVO_TRACA
98.2 Ovomucoid P67946 IOVO_TRABL
98.2 Ovomucoid P67960 IOVO_SYRSO
98.2 Ovomucoid P67959 IOVO_SYRHU
98.2 Ovomucoid P67958 IOVO_SYREL
98.2 Ovomucoid P67956 IOVO_PUCMA
98.2 Ovomucoid P67955 IOVO_LOPSW
98.2 Ovomucoid P67954 IOVO_LOPNY
98.2 Ovomucoid P67950 IOVO_LOPIM
98.2 Ovomucoid P67953 IOVO_LOPED
98.2 Ovomucoid P67961 IOVO_LAGLU
98.2 Ovomucoid P67952 IOVO_CROMA
98.2 Ovomucoid P67951 IOVO_CROAU
98.2 Ovomucoid P67945 IOVO_CENUR
98.2 Ovomucoid P67957 IOVO_CATWA
100.0 Ovomucoid P68436 IOVO_MELOE
100.0 Ovomucoid P68390 IOVO_MELGA
100.0 Ovomucoid P52245 IOVO_CROCS
91.1 Ovomucoid P52259 IOVO_BAMTH