The native state conformational ensemble of the SH3 domain from alpha-spectrin.


The folding/unfolding equilibrium of the alpha-spectrin SH3 domain has been measured by NMR-detected hydrogen/deuterium exchange and by differential scanning calorimetry. Protection factors against exchange have been obtained under native conditions for more than half of the residues in the domain. Most protected residues are located at the beta-strands, the short 3(10) helix, and part of the long RT loop, whereas the loops connecting secondary structure elements show no measurable protection. Apparent stability constants per residue and their corresponding Gibbs energies have been calculated from the exchange experiments. The most stable region of the SH3 domain is defined by the central portions of the beta-strands. The peptide binding region, on the other hand, is composed of a highly stable region (residues 53-57) and a highly unstable region, the loop between residues 34-41 (n-Src loop). All residues in the domain have apparent Gibbs energies lower than the global unfolding Gibbs energy measured by differential scanning calorimetry, indicating that under our experimental conditions the amide exchange of all residues in the SH3 domain occurs primarily via local unfolding reactions. A structure-based thermodynamic analysis has allowed us to predict correctly the thermodynamics of the global unfolding of the domain and to define the ensemble of conformational states that quantitatively accounts for the observed pattern of hydrogen exchange protection. These results demonstrate that under native conditions the SH3 domain needs to be considered as an ensemble of conformations and that the hydrogen exchange data obtained under those conditions cannot be interpreted by a two-state equilibrium. The observation that specific regions of a protein are able to undergo independent local folding/unfolding reactions indicates that under native conditions the scale of cooperative interactions is regional rather than global. Study holds ProTherm entries: 23822, 23823, 23824, 23825, 23826, 23827, 23828, 23829, 23830, 23831, 23832, 23833, 23834 Extra Details: folding/unfolding equilibrium, alpha-spectrin SH3 domain, differential scanning calorimetry.

Submission Details

ID: fEawmRs73

Submitter: Connie Wang

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

Version: 1

Publication Details
Sadqi M;Casares S;Abril MA;López-Mayorga O;Conejero-Lara F;Freire E,Biochemistry (1999) The native state conformational ensemble of the SH3 domain from alpha-spectrin. PMID:10413463
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 UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
100.0 Spectrin alpha chain, non-erythrocytic 1 P07751 SPTN1_CHICK
100.0 Spectrin alpha chain, non-erythrocytic 1 Q13813 SPTN1_HUMAN
100.0 Spectrin alpha chain, non-erythrocytic 1 P16546 SPTN1_MOUSE
100.0 Spectrin alpha chain, non-erythrocytic 1 P16086 SPTN1_RAT