Thermodynamic analysis of the folding of the streptococcal protein G IgG-binding domains B1 and B2: why small proteins tend to have high denaturation temperatures.


Abstract

We have cloned, expressed, and characterized two naturally occurring variations of the IgG-binding domain of streptococcal protein G. The domain is a stable cooperative folding unit of 56 amino acids, which maintains a unique folded structure without disulfide cross-links or tight ligand binding. We have studied the thermodynamics of the unfolding reaction for the two versions of this domain, designated B1 and B2, which differ by six amino acids. They have denaturation temperatures of 87.5 degrees C and 79.4 degrees C, respectively at pH 5.4, as determined by differential scanning calorimetry. Thermodynamic state functions for the unfolding reaction (delta G, delta H, delta S, and delta Cp) have been determined and reveal several interesting insights into the behavior of very small proteins. First, though the B1 domain has a heat denaturation point close to 90 degrees C, it is not unusually stable at physiologically relevant temperatures (delta G = 25 kJ/mol at 37 degrees C). This behavior occurs because the stability profile (delta G vs temperature) is flat and shallow due to the small delta S and delta Cp for unfolding. Related to this point is the second observation that small changes in the free energy of unfolding of the B-domain due to mutation or change in solvent conditions lead to large shifts in the heat denaturation temperature. Third, the magnitude and relative contributions of hydrophobic vs nonhydrophobic forces (per amino acid residue) to the total free energy of folding of the B-domain are remarkably typical of other globular proteins of much larger size. Study holds ProTherm entries: 2888, 2889, 2890, 2891, 2892, 2893, 2894, 2895 Extra Details: streptococcus protein G; IgG-binding domains; folding;,thermodynamic analysis; hydrophobic

Submission Details

ID: b5TToFjA

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:20 p.m.

Version: 1

Publication Details
Alexander P;Fahnestock S;Lee T;Orban J;Bryan P,Biochemistry (1992) Thermodynamic analysis of the folding of the streptococcal protein G IgG-binding domains B1 and B2: why small proteins tend to have high denaturation temperatures. PMID:1567818
Additional Information

Study Summary

Number of data points 32
Proteins Immunoglobulin G-binding protein G ; Immunoglobulin G-binding protein G
Unique complexes 1
Assays/Quantities/Protocols Experimental Assay: dCp buffers:glycine: 50 mM, pH:5.4 ; Experimental Assay: dHcal buffers:glycine: 50 mM, pH:5.4 ; Experimental Assay: Tm buffers:glycine: 50 mM, pH:5.4 ; Experimental Assay: dHvH buffers:glycine: 50 mM, pH:5.4 ; Experimental Assay: dCp pH:4.0, buffers:glycine: 50 mM ; Experimental Assay: dHcal pH:4.0, buffers:glycine: 50 mM ; Experimental Assay: Tm pH:4.0, buffers:glycine: 50 mM ; Experimental Assay: dHvH pH:4.0, buffers:glycine: 50 mM ; Experimental Assay: dCp pH:3.1, buffers:glycine: 50 mM ; Experimental Assay: dHcal pH:3.1, buffers:glycine: 50 mM ; Experimental Assay: Tm pH:3.1, buffers:glycine: 50 mM ; Experimental Assay: dHvH pH:3.1, buffers:glycine: 50 mM ; Experimental Assay: dCp buffers:glycine: 50 mM, pH:2.88 ; Experimental Assay: dHcal buffers:glycine: 50 mM, pH:2.88 ; Experimental Assay: Tm buffers:glycine: 50 mM, pH:2.88 ; Experimental Assay: dHvH buffers:glycine: 50 mM, pH:2.88 ; Experimental Assay: dCp pH:2.69, buffers:glycine: 50 mM ; Experimental Assay: dHcal pH:2.69, buffers:glycine: 50 mM ; Experimental Assay: Tm pH:2.69, buffers:glycine: 50 mM ; Experimental Assay: dHvH pH:2.69, buffers:glycine: 50 mM ; Experimental Assay: dCp buffers:Sodium acetate: 50 mM, pH:5.4 ; Experimental Assay: dHcal buffers:Sodium acetate: 50 mM, pH:5.4 ; Experimental Assay: Tm buffers:Sodium acetate: 50 mM, pH:5.4 ; Experimental Assay: dHvH buffers:Sodium acetate: 50 mM, pH:5.4 ; Experimental Assay: dCp pH:3.51, buffers:Sodium acetate: 50 mM ; Experimental Assay: dHcal pH:3.51, buffers:Sodium acetate: 50 mM ; Experimental Assay: Tm pH:3.51, buffers:Sodium acetate: 50 mM ; Experimental Assay: dHvH pH:3.51, buffers:Sodium acetate: 50 mM ; Experimental Assay: dCp pH:2.3, buffers:Sodium acetate: 50 mM ; Experimental Assay: dHcal pH:2.3, buffers:Sodium acetate: 50 mM ; Experimental Assay: Tm pH:2.3, buffers:Sodium acetate: 50 mM ; Experimental Assay: dHvH pH:2.3, buffers:Sodium acetate: 50 mM
Libraries Mutations for sequence MTYKLILNGKTLKGETTTEAVDAATAEKVFKQYANDNGVDGEWTYDDATKTFTVTE

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
1EM7 2000-03-16T00:00:00+0000 2.0 HELIX VARIANT OF THE B1 DOMAIN FROM STREPTOCOCCAL PROTEIN G
1GB1 1991-05-15T00:00:00+0000 0 A NOVEL, HIGHLY STABLE FOLD OF THE IMMUNOGLOBULIN BINDING DOMAIN OF STREPTOCOCCAL PROTEIN G
1IGC 1994-08-05T00:00:00+0000 2.6 IGG1 FAB FRAGMENT (MOPC21) COMPLEX WITH DOMAIN III OF PROTEIN G FROM STREPTOCOCCUS
1IGD 1994-08-05T00:00:00+0000 1.1 THE THIRD IGG-BINDING DOMAIN FROM STREPTOCOCCAL PROTEIN G: AN ANALYSIS BY X-RAY CRYSTALLOGRAPHY OF THE STRUCTURE ALONE AND IN A COMPLEX WITH FAB
1LE3 2002-04-09T00:00:00+0000 0 NMR Structure of Tryptophan Zipper 4: A Stable Beta-Hairpin Peptide Based on the C-terminal Hairpin of the B1 Domain of Protein G
1MPE 2002-09-12T00:00:00+0000 0 Ensemble of 20 structures of the tetrameric mutant of the B1 domain of streptococcal protein G
1MVK 2002-09-25T00:00:00+0000 2.5 X-ray structure of the tetrameric mutant of the B1 domain of streptococcal protein G
1PGA 1993-11-23T00:00:00+0000 2.07 TWO CRYSTAL STRUCTURES OF THE B1 IMMUNOGLOBULIN-BINDING DOMAIN OF STREPTOCOCCAL PROTEIN G AND COMPARISON WITH NMR
1PGB 1993-11-23T00:00:00+0000 1.92 TWO CRYSTAL STRUCTURES OF THE B1 IMMUNOGLOBULIN-BINDING DOMAIN OF STREPTOCCOCAL PROTEIN G AND COMPARISON WITH NMR
1PGX 1992-04-03T00:00:00+0000 1.66 THE 1.66 ANGSTROMS X-RAY STRUCTURE OF THE B2 IMMUNOGLOBULIN-BINDING DOMAIN OF STREPTOCOCCAL PROTEIN G AND COMPARISON TO THE NMR STRUCTURE OF THE B1 DOMAIN

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
100.0 Immunoglobulin G-binding protein G P19909 SPG2_STRSG
100.0 Immunoglobulin G-binding protein G P06654 SPG1_STRSG