Contribution of salt bridges near the surface of a protein to the conformational stability.


Abstract

Salt bridges play important roles in the conformational stability of proteins. However, the effect of a surface salt bridge on the stability remains controversial even today; some reports have shown little contribution of a surface salt bridge to stability, whereas others have shown a favorable contribution. In this study, to elucidate the net contribution of a surface salt bridge to the conformational stability of a protein, systematic mutant human lysozymes, containing one Glu to Gln (E7Q) and five Asp to Asn mutations (D18N, D49N, D67N, D102N, and D120N) at residues where a salt bridge is formed near the surface in the wild-type structure, were examined. The thermodynamic parameters for denaturation between pH 2.0 and 4.8 were determined by use of a differential scanning calorimeter, and the crystal structures were analyzed by X-ray crystallography. The denaturation Gibbs energy (DeltaG) of all mutant proteins was lower than that of the wild-type protein at pH 4, whereas there was little difference between them near pH 2. This is caused by the fact that the Glu and Asp residues are ionized at pH 4 but protonated at pH 2, indicating a favorable contribution of salt bridges to the wild-type structure at pH 4. Each contribution was not equivalent, but we found that the contributions correlate with the solvent inaccessibility of the salt bridges; the salt bridge contribution was small when 100% accessible, while it was about 9 kJ/mol if 100% inaccessible. This conclusion indicates how to reconcile a number of conflicting reports about role of surface salt bridges in protein stability. Furthermore, the effect of salts on surface salt bridges was also examined. In the presence of 0.2 M KCl, the stability at pH 4 decreased, and the differences in stability between the wild-type and mutant proteins were smaller than those in the absence of salts, indicating the compensation to the contribution of salt bridges with salts. Salt bridges with more than 50% accessibility did not contribute to the stability in the presence of 0.2 M KCl. Study holds ProTherm entries: 10238, 10239, 10240, 10241, 10242, 10243, 10244, 10245, 10246, 10247, 10248, 10249, 10250, 10251, 10266, 10267, 10268, 10269, 10270, 10271, 10272, 10273, 10274, 10275, 14540, 14541, 14542, 14543, 14544, 14545, 14546, 14547, 14548, 14549, 14550, 14551, 14552, 14553, 14554, 14555, 14556, 14557, 14558, 14559 Extra Details: salt bridges; conformational stability; thermodynamic parameters;,solvent inaccessibility

Submission Details

ID: HLF69ceE4

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:40 p.m.

Version: 1

Publication Details
Takano K;Tsuchimori K;Yamagata Y;Yutani K,Biochemistry (2000) Contribution of salt bridges near the surface of a protein to the conformational stability. PMID:11015217
Additional Information

Study Summary

Number of data points 84
Proteins Lysozyme C ; Lysozyme C
Unique complexes 7
Assays/Quantities/Protocols Experimental Assay: dG pH:4.5, buffers:sodium acetate: 0.02 M, ionic:KCl: 0.2 M ; Experimental Assay: dG pH:2.3, ionic:KCl: 0.2 M, buffers:glycine hydrochloride: 0.05 M ; Experimental Assay: dG buffers:sodium acetate: 0.02 M, ionic:-: -, pH:4.0 ; Experimental Assay: dG ionic:-: -, pH:2.2, buffers:glycine hydrochloride: 0.05 M ; Experimental Assay: Tm pH:4.5, buffers:sodium acetate: 0.02 M, ionic:KCl: 0.2 M ; Experimental Assay: Tm pH:2.3, ionic:KCl: 0.2 M, buffers:glycine hydrochloride: 0.05 M ; Experimental Assay: Tm pH:4.0, ionic:: , buffers:sodium acetate: 0.02 M ; Experimental Assay: Tm ionic:: , pH:2.2, buffers:glycine hydrochloride: 0.05 M ; Derived Quantity: ddG pH:4.5, buffers:sodium acetate: 0.02 M, ionic:KCl: 0.2 M ; Derived Quantity: ddG pH:2.3, ionic:KCl: 0.2 M, buffers:glycine hydrochloride: 0.05 M ; Derived Quantity: ddG buffers:sodium acetate: 0.02 M, ionic:-: -, pH:4.0 ; Derived Quantity: ddG ionic:-: -, pH:2.2, buffers:glycine hydrochloride: 0.05 M ; Derived Quantity: dTm pH:4.5, buffers:sodium acetate: 0.02 M, ionic:KCl: 0.2 M ; Derived Quantity: dTm pH:2.3, ionic:KCl: 0.2 M, buffers:glycine hydrochloride: 0.05 M ; Derived Quantity: dTm pH:4.0, ionic:: , buffers:sodium acetate: 0.02 M ; Derived Quantity: dTm ionic:: , pH:2.2, buffers:glycine hydrochloride: 0.05 M
Libraries Mutations for sequence KVFERCELARTLKRLGMDGYRGISLANWMCLAKWESGYNTRATNYNAGDRSTDYGIFQINSRYWCNDGKTPGAVNACHLSCSALLQDNIADAVACAKRVVRDPQGIRAWVAWRNRCQNRDVRQYVQGCGV

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
133L 1993-06-01T00:00:00+0000 1.77 ROLE OF ARG 115 IN THE CATALYTIC ACTION OF HUMAN LYSOZYME. X-RAY STRUCTURE OF HIS 115 AND GLU 115 MUTANTS
134L 1993-06-01T00:00:00+0000 1.77 ROLE OF ARG 115 IN THE CATALYTIC ACTION OF HUMAN LYSOZYME. X-RAY STRUCTURE OF HIS 115 AND GLU 115 MUTANTS
1B5U 1999-01-11T00:00:00+0000 1.8 CONTRIBUTION OF HYDROGEN BONDS TO THE CONFORMATIONAL STABILITY OF HUMAN LYSOZYME: CALORIMETRY AND X-RAY ANALYSIS OF SIX SER->ALA MUTANT
1B5V 1999-01-11T00:00:00+0000 2.17 CONTRIBUTION OF HYDROGEN BONDS TO THE CONFORMATIONAL STABILITY OF HUMAN LYSOZYME: CALORIMETRY AND X-RAY ANALYSIS OF SIX SER->ALA MUTANTS
1B5W 1999-01-11T00:00:00+0000 2.17 CONTRIBUTION OF HYDROGEN BONDS TO THE CONFORMATIONAL STABILITY OF HUMAN LYSOZYME: CALORIMETRY AND X-RAY ANALYSIS OF SIX SER->ALA MUTANTS
1B5X 1999-01-11T00:00:00+0000 2.0 Contribution of hydrogen bonds to the conformational stability of human lysozyme: calorimetry and x-ray analysis of six ser->ala mutants
1B5Y 1999-01-11T00:00:00+0000 2.2 CONTRIBUTION OF HYDROGEN BONDS TO THE CONFORMATIONAL STABILITY OF HUMAN LYSOZYME: CALORIMETRY AND X-RAY ANALYSIS OF SIX SER->ALA MUTANTS
1B5Z 1999-01-11T00:00:00+0000 2.2 CONTRIBUTION OF HYDROGEN BONDS TO THE CONFORMATIONAL STABILITY OF HUMAN LYSOZYME: CALORIMETRY AND X-RAY ANALYSIS OF SIX SER->ALA MUTANTS
1B7L 1999-01-24T00:00:00+0000 1.8 VERIFICATION OF SPMP USING MUTANT HUMAN LYSOZYMES
1B7M 1999-01-24T00:00:00+0000 2.2 VERIFICATION OF SPMP USING MUTANT HUMAN LYSOZYMES

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
100.0 Lysozyme C P61626 LYSC_HUMAN
100.0 Lysozyme C P61627 LYSC_PANPA
100.0 Lysozyme C P61628 LYSC_PANTR
100.0 Lysozyme C P79179 LYSC_GORGO
99.2 Lysozyme C P79239 LYSC_PONPY
96.9 Lysozyme C P79180 LYSC_HYLLA