Folding subdomains of thioredoxin characterized by native-state hydrogen exchange.


Native-state hydrogen exchange (HX) studies, used in conjunction with NMR spectroscopy, have been carried out on Escherichia coli thioredoxin (Trx) for characterizing two folding subdomains of the protein. The backbone amide protons of only the slowest-exchanging 24 amino acid residues, of a total of 108 amino acid residues, could be followed at pH 7. The free energy of the opening event that results in an amide hydrogen exchanging with solvent (DeltaG(op)) was determined at each of the 24 amide hydrogen sites. The values of DeltaG(op) for the amide hydrogens belonging to residues in the helices alpha(1), alpha(2), and alpha(4) are consistent with them exchanging with the solvent only when the fully unfolded state is sampled transiently under native conditions. The denaturant-dependences of the values of DeltaG(op) provide very little evidence that the protein samples partially unfolded forms, lower in energy than the unfolded state. The amide hydrogens belonging to the residues in the beta strands, which form the core of the protein, appear to have higher values of DeltaG(op) than amide hydrogens belonging to residues in the helices, suggesting that they might be more stable to exchange. This apparently higher stability to HX of the beta strands might be either because they exchange out their amide hydrogens in a high energy intermediate preceding the globally unfolded state, or, more likely, because they form residual structure in the globally unfolded state. In either case, the central beta strands-beta(3,) beta(2), and beta(4)-would appear to form a cooperatively folding subunit of the protein. The native-state HX methodology has made it possible to characterize the free energy landscape that Trx can sample under equilibrium native conditions. Study holds ProTherm entries: 16585, 16586 Extra Details: native-state HX; NMR spectroscopy; thioredoxin; folding; unfolded state

Submission Details

ID: xb6Xh6BM4

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:48 p.m.

Version: 1

Publication Details
Bhutani N;Udgaonkar JB,Protein Sci. (2003) Folding subdomains of thioredoxin characterized by native-state hydrogen exchange. PMID:12876321
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
3DXB 2008-07-24T00:00:00+0000 2.2 Structure of the UHM domain of Puf60 fused to thioredoxin
5E4W 2015-10-07T00:00:00+0000 2.8 Crystal structure of cpSRP43 chromodomains 2 and 3 in complex with the Alb3 tail
5IKN 2016-03-03T00:00:00+0000 4.8 Crystal Structure of the T7 Replisome in the Absence of DNA
1KEB 2001-11-15T00:00:00+0000 1.8 Crystal Structure of Double Mutant M37L,P40S E.coli Thioredoxin
1M7T 2002-07-22T00:00:00+0000 0 Solution Structure and Dynamics of the Human-Escherichia coli Thioredoxin Chimera: Insights into Thermodynamic Stability
1OAZ 2003-01-21T00:00:00+0000 2.78 IgE Fv SPE7 complexed with a recombinant thioredoxin
1SKR 2004-03-05T00:00:00+0000 2.4 T7 DNA Polymerase Complexed To DNA Primer/Template and ddATP
1SKS 2004-03-05T00:00:00+0000 2.3 Binary 3' complex of T7 DNA polymerase with a DNA primer/template containing a cis-syn thymine dimer on the template
1SKW 2004-03-05T00:00:00+0000 2.3 Binary 3' complex of T7 DNA polymerase with a DNA primer/template containing a disordered cis-syn thymine dimer on the template

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
100.0 Thioredoxin 1 P0AA30 THIO_SHIFL
100.0 Thioredoxin 1 P0AA28 THIO_SALTY
100.0 Thioredoxin 1 P0AA29 THIO_SALTI
100.0 Thioredoxin 1 P0AA25 THIO_ECOLI
100.0 Thioredoxin 1 P0AA26 THIO_ECOL6
100.0 Thioredoxin 1 P0AA27 THIO_ECO57