Probing the structure and stability of a hybrid protein: the human-E. coli thioredoxin chimera.


Abstract

The structure and stability of a hybrid protein composed of N-terminal human and C-terminal E. coli thioredoxin domains were investigated by NMR, fluorescence, and circular dichroism spectroscopy. We demonstrate that the chimeric protein is correctly folded and exhibits the common thioredoxin architecture. However, the stability of the hybrid protein toward thermal and chemical denaturation is clearly reduced when compared with both parent proteins. Altogether, our data indicate that the interface between the two folding units of thioredoxin is tolerant toward changes in exact interdigitation of side chains, allowing for the formation of the unique overall thioredoxin fold. Further, the gene encoding the human-E. coli chimera was tested in vivo whether it supports the assembly of filamentous phages. No complementation of a thioredoxin-deficient E. coli mutant for the replication of the phages M13 or fd was observed, suggesting that parts of the overall protein structure in the N-terminal domain are crucial for this activity. Study holds ProTherm entries: 11712, 11713 Extra Details: chimeric protein; hybrid; thioredoxin fold; activity

Submission Details

ID: jwvKeGNf

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:42 p.m.

Version: 1

Publication Details
Louis JM;Georgescu RE;Tasayco ML;Tcherkasskaya O;Gronenborn AM,Biochemistry (2001) Probing the structure and stability of a hybrid protein: the human-E. coli thioredoxin chimera. PMID:11551217
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
1AIU 1997-04-25T00:00:00+0000 2.0 HUMAN THIOREDOXIN (D60N MUTANT, REDUCED FORM)
1AUC 1997-08-22T00:00:00+0000 2.1 HUMAN THIOREDOXIN (OXIDIZED WITH DIAMIDE)
1CQG 1996-04-02T00:00:00+0000 0 HIGH RESOLUTION SOLUTION NMR STRUCTURE OF MIXED DISULFIDE INTERMEDIATE BETWEEN HUMAN THIOREDOXIN (C35A, C62A, C69A, C73A) MUTANT AND A 13 RESIDUE PEPTIDE COMPRISING ITS TARGET SITE IN HUMAN REF-1 (RESIDUES 59-71 OF THE P50 SUBUNIT OF NFKB), NMR, 31 STRUCTURES
1CQH 1996-04-02T00:00:00+0000 0 HIGH RESOLUTION SOLUTION NMR STRUCTURE OF MIXED DISULFIDE INTERMEDIATE BETWEEN HUMAN THIOREDOXIN (C35A, C62A, C69A, C73A) MUTANT AND A 13 RESIDUE PEPTIDE COMPRISING ITS TARGET SITE IN HUMAN REF-1 (RESIDUES 59-71 OF THE P50 SUBUNIT OF NFKB), NMR, MINIMIZED AVERAGE STRUCTURE
1ERT 1996-02-07T00:00:00+0000 1.7 HUMAN THIOREDOXIN (REDUCED FORM)
1ERU 1996-02-07T00:00:00+0000 2.1 HUMAN THIOREDOXIN (OXIDIZED FORM)
1ERV 1996-02-07T00:00:00+0000 1.65 HUMAN THIOREDOXIN MUTANT WITH CYS 73 REPLACED BY SER (REDUCED FORM)

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
90.3 Thioredoxin P11232 THIO_RAT
90.5 Thioredoxin O97508 THIO_HORSE
92.4 Thioredoxin P50413 THIO_SHEEP
93.3 Thioredoxin O97680 THIO_BOVIN
94.3 Thioredoxin P82460 THIO_PIG
96.2 Thioredoxin Q9BDJ3 THIO_CALJA
99.1 Thioredoxin Q5R9M3 THIO_PONAB
97.1 Thioredoxin P29451 THIO_MACMU
100.0 Thioredoxin P10599 THIO_HUMAN