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


Abstract

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 PDB Entries

Structure ID Release Date Resolution Structure Title
3THK 2011-08-19T00:00:00+0000 1.7 Structure of SH3 chimera with a type II ligand linked to the chain C-terminal
3FB2 2008-11-18T00:00:00+0000 2.3 Crystal structure of the human brain alpha spectrin repeats 15 and 16. Northeast Structural Genomics Consortium target HR5563a.
6ZEH 2020-06-16T00:00:00+0000 1.3 Structure of PP1-spectrin alpha II chimera [PP1(7-304) + linker (G/S)x9 + spectrin alpha II (1025-1039)] bound to Phactr1 (516-580)
2FOT 2006-01-13T00:00:00+0000 2.45 Crystal structure of the complex between calmodulin and alphaII-spectrin
3F31 2008-10-30T00:00:00+0000 2.3 Crystal Structure of the N-terminal region of AlphaII-spectrin Tetramerization Domain
5FW9 2016-02-12T00:00:00+0000 1.55 Human Spectrin SH3 domain D48G, E7Y, K60Y
5FWC 2016-02-12T00:00:00+0000 1.4 Human Spectrin SH3 domain D48G, E7A, K60A
5FWB 2016-02-12T00:00:00+0000 1.5 Human Spectrin SH3 domain D48G, E7F, K60F
1SHG 1993-05-19T00:00:00+0000 1.8 CRYSTAL STRUCTURE OF A SRC-HOMOLOGY 3 (SH3) DOMAIN
2JMA 2006-10-25T00:00:00+0000 0 R21A Spc-SH3:P41 complex

Relevant UniProtKB Entries

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