Determining stability of proteins from guanidinium chloride transition curves.


Abstract

The guanidinium chloride (GdmCl) denaturation of RNAase A, lysozyme and metmyoglobin was investigated at several pH values by using absorbance measurements at 287, 300 and 409 nm respectively. From these measurements the free-energy change on denaturation, delta Gapp., was calculated, assuming a two-state mechanism, and values of delta Gapp. at zero concentration of the denaturant were measured. For each protein all delta Gapp. values were adjusted to pH 7.00 by using the appropriate relationship between delta Gapp. and pH. Dependence of the adjusted delta Gapp. value on GdmCl concentration increases for metmyoglobin and decreases for the other two proteins as the denaturant concentration decreases. It has been shown that these are expected results if the presence of the acid-denatured state during the GdmCl denaturation of proteins is considered. Study holds ProTherm entries: 7718, 7719, 7720, 7721, 7722, 7723, 7724, 7725, 7726, 7727, 7728, 7729, 7730, 7731, 7732 Extra Details: guanidinium chloride transition curve; absorbance measurement;,free-energy change; two-state mechanism; acid-denatured state

Submission Details

ID: mX63tSLC3

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:34 p.m.

Version: 1

Publication Details
Ahmad F;Yadav S;Taneja S,Biochem. J. (1992) Determining stability of proteins from guanidinium chloride transition curves. PMID:1445206
Additional Information

Study Summary

Number of data points 45
Proteins Ribonuclease pancreatic ; Myoglobin ; Lysozyme C ; Ribonuclease pancreatic ; Myoglobin ; Lysozyme C
Unique complexes 3
Assays/Quantities/Protocols Experimental Assay: Cm pH:4.6 ; Experimental Assay: m pH:4.6 ; Experimental Assay: dG_H2O pH:4.6 ; Experimental Assay: Cm pH:4.92 ; Experimental Assay: m pH:4.92 ; Experimental Assay: dG_H2O pH:4.92 ; Experimental Assay: Cm pH:5.26 ; Experimental Assay: m pH:5.26 ; Experimental Assay: dG_H2O pH:5.26 ; Experimental Assay: Cm pH:5.46 ; Experimental Assay: m pH:5.46 ; Experimental Assay: dG_H2O pH:5.46 ; Experimental Assay: Cm pH:5.52 ; Experimental Assay: m pH:5.52 ; Experimental Assay: dG_H2O pH:5.52 ; Experimental Assay: Cm pH:5.88 ; Experimental Assay: m pH:5.88 ; Experimental Assay: dG_H2O pH:5.88 ; Experimental Assay: Cm pH:0.34 ; Experimental Assay: m pH:0.34 ; Experimental Assay: dG_H2O pH:0.34 ; Experimental Assay: Cm pH:0.64 ; Experimental Assay: m pH:0.64 ; Experimental Assay: dG_H2O pH:0.64 ; Experimental Assay: Cm pH:2.03 ; Experimental Assay: m pH:2.03 ; Experimental Assay: dG_H2O pH:2.03 ; Experimental Assay: Cm pH:6.0 ; Experimental Assay: m pH:6.0 ; Experimental Assay: dG_H2O pH:6.0 ; Experimental Assay: Cm pH:1.3 ; Experimental Assay: m pH:1.3 ; Experimental Assay: dG_H2O pH:1.3 ; Experimental Assay: Cm pH:2.1 ; Experimental Assay: m pH:2.1 ; Experimental Assay: dG_H2O pH:2.1 ; Experimental Assay: Cm pH:3.0 ; Experimental Assay: m pH:3.0 ; Experimental Assay: dG_H2O pH:3.0 ; Experimental Assay: Cm pH:7.0 ; Experimental Assay: m pH:7.0 ; Experimental Assay: dG_H2O pH:7.0
Libraries Mutations for sequence KVFGRCELAAAMKRHGLDNYRGYSLGNWVCAAKFESNFNTQATNRNTDGSTDYGILQINSRWWCNDGRTPGSRNLCNIPCSALLSSDITASVNCAKKIVSDGNGMNAWVAWRNRCKGTDVQAWIRGCRL ; Mutations for sequence GLSDGEWQQVLNVWGKVEADIAGHGQEVLIRLFTGHPETLEKFDKFKHLKTEAEMKASEDLKKHGTVVLTALGGILKKKGHHEAELKPLAQSHATKHKIPIKYLEFISDAIIHVLHSKHPGDFGADAQGAMTKALELFRNDIAAKYKELGFQG ; Mutations for sequence KETAAAKFERQHMDSSTSAASSSNYCNQMMKSRNLTKDRCKPVNTFVHESLADVQAVCSQKNVACKNGQTNCYQSYSTMSITDCRETGSSKYPNCAYKTTQANKHIIVACEGNPYVPVHFDASV

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
2IHL 1993-06-29T00:00:00+0000 1.4 LYSOZYME (E.C.3.2.1.17) (JAPANESE QUAIL)
6QMN 2019-02-07T00:00:00+0000 2.31 Crystal structure of a Ribonuclease A-Onconase chimera
1FBI 1995-01-19T00:00:00+0000 3.0 CRYSTAL STRUCTURE OF A CROSS-REACTION COMPLEX BETWEEN FAB F9.13.7 AND GUINEA-FOWL LYSOZYME
1GHL 1993-05-04T00:00:00+0000 2.1 THE THREE-DIMENSIONAL STRUCTURE OF PHEASANT AND GUINEA-FOWL EGG LYSOZYMES
1HHL 1993-05-04T00:00:00+0000 1.9 THE THREE-DIMENSIONAL STRUCTURE OF PHEASANT AND GUINEA-FOWL EGG LYSOZYMES
1JHL 1993-05-04T00:00:00+0000 2.4 THREE-DIMENSIONAL STRUCTURE OF A HETEROCLITIC ANTIGEN-ANTIBODY CROSS-REACTION COMPLEX
5YCI 2017-09-07T00:00:00+0000 1.97 Ancestral myoglobin aMbWb' of Basilosaurus relative (polyphyly)
5YCJ 2017-09-07T00:00:00+0000 1.58 Ancestral myoglobin aMbWb' of Basilosaurus relative (polyphyly) imidazole-ligand
1BQL 1995-02-03T00:00:00+0000 2.6 STRUCTURE OF AN ANTI-HEL FAB FRAGMENT COMPLEXED WITH BOBWHITE QUAIL LYSOZYME
1DKJ 1996-01-10T00:00:00+0000 2.0 BOBWHITE QUAIL LYSOZYME

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
96.0 Ribonuclease pancreatic Q29606 RNAS1_ORYLE
90.3 Ribonuclease pancreatic P00662 RNAS1_GIRCA
92.7 Ribonuclease pancreatic P00668 RNAS1_ANTAM
95.2 Ribonuclease pancreatic P00660 RNAS1_CONTA
93.5 Ribonuclease pancreatic P07848 RNAS1_EUDTH
96.0 Ribonuclease pancreatic P07847 RNAS1_AEPME
95.2 Ribonuclease pancreatic P00657 RNAS1_BUBBU
96.8 Ribonuclease pancreatic P67927 RNAS1_SHEEP
96.8 Ribonuclease pancreatic P67926 RNAS1_CAPHI
100.0 Ribonuclease pancreatic P61824 RNAS1_BISBI
100.0 Ribonuclease pancreatic P61823 RNAS1_BOVIN
90.3 Myoglobin P02163 MYG_ROUAE
90.9 Myoglobin P11343 MYG_LUTLU
90.3 Myoglobin P02165 MYG_TUPGL
90.3 Myoglobin P02167 MYG_NYCCO
90.3 Myoglobin Q0KIY0 MYG_MESST
90.3 Myoglobin P02183 MYG_MESCA
90.8 Myoglobin P02177 MYG_ESCRO
90.8 Myoglobin Q0KIY2 MYG_BALED
90.8 Myoglobin Q0KIY1 MYG_BALBO
90.9 Myoglobin P02189 MYG_PIG
90.9 Myoglobin P02166 MYG_PERPO
92.2 Myoglobin P02169 MYG_LEPMU
91.6 Myoglobin P02181 MYG_INIGE
100.0 Myoglobin P68082 MYG_HORSE
100.0 Myoglobin P68083 MYG_EQUBU
93.0 Lysozyme C P81711 LYSC_SYRSO
92.3 Lysozyme C P49663 LYSC_PHAVE
94.6 Lysozyme C P24533 LYSC_SYRRE
93.0 Lysozyme C P24364 LYSC_LOPLE
92.2 Lysozyme C P00704 LYSC_NUMME
95.3 Lysozyme C Q7LZT2 LYSC_TRATE
95.3 Lysozyme C P22910 LYSC_CHRAM
96.1 Lysozyme C P19849 LYSC_PAVCR
96.1 Lysozyme C Q7LZI3 LYSC_TRASA
96.9 Lysozyme C Q7LZP9 LYSC_LOPIM
96.9 Lysozyme C Q7LZQ0 LYSC_CATWA
96.9 Lysozyme C P00699 LYSC_CALCC
96.9 Lysozyme C P00700 LYSC_COLVI
93.2 Lysozyme C P00702 LYSC_PHACO
95.2 Lysozyme C P00703 LYSC_MELGA
95.3 Lysozyme C P00701 LYSC_COTJA
100.0 Lysozyme C P00698 LYSC_CHICK
90.1 Myoglobin P02178 MYG_MEGNO