The reactive and destabilizing disulfide bond of DsbA, a protein required for protein disulfide bond formation in vivo.


Abstract

The protein DsbA facilitates disulfide bond formation in the periplasm of Escherichia coli. It has only two cysteine residues that are separated in the sequence by two other residues and are shown to form a disulfide bond reversibly. Chemical modification studies demonstrate that only one of the cysteine residues has an accessible thiol group in the reduced protein. Equilibrium and kinetic characterization of thiol-disulfide exchange between DsbA and glutathione showed that the DsbA disulfide bond was 10(3)-fold more reactive than a normal protein disulfide. Similarly, the mixed disulfide between the accessible cysteine residue and glutathione was 10(4)-fold more reactive than normal. The overall equilibrium constant for DsbA disulfide bond formation from GSSG was only 8 x 10(-5) M. These properties indicate that disulfide-bonded DsbA is a potent oxidant and ideally suited for generating protein disulfide bonds. Disulfide bonds generally increase the stabilities of folded proteins, when the folded conformation reciprocally stabilizes the disulfide bonds. In contrast, the disulfide bond of DsbA was so unstable in the folded state that its stability increased by 4.5 +/- 0.1 kcal.mol-1 when the protein unfolded. This implies that the disulfide bond destabilizes the folded conformation of DsbA. This was confirmed by demonstrating that the reduced protein was 3.6 +/- 1.4 kcal.mol-1 more stable than that with the disulfide bond. Study holds ProTherm entries: 4572, 4573 Extra Details: additive : EDTA(1 mM), disulfide bond formation; cysteine; ccessible;,folded conformation

Submission Details

ID: gV6M8FCu3

Submitter: Connie Wang

Submission Date: April 24, 2018, 8:26 p.m.

Version: 1

Publication Details
Zapun A;Bardwell JC;Creighton TE,Biochemistry (1993) The reactive and destabilizing disulfide bond of DsbA, a protein required for protein disulfide bond formation in vivo. PMID:8494885
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
2LEG 2011-10-26 Membrane protein complex DsbB-DsbA structure by joint calculations with solid-state NMR and X-ray experimental data
2NDO 2017-02-08 Structure of EcDsbA-sulfonamide1 complex
1A24 1998-09-16 SOLUTION NMR STRUCTURE OF REDUCED DSBA FROM ESCHERICHIA COLI, FAMILY OF 20 STRUCTURES
1A23 1998-09-16 SOLUTION NMR STRUCTURE OF REDUCED DSBA FROM ESCHERICHIA COLI, MINIMIZED AVERAGE STRUCTURE
1FVK 1997-08-20 1.7 THE 1.7 ANGSTROM STRUCTURE OF WILD TYPE DISULFIDE BOND FORMATION PROTEIN (DSBA)
4ZIJ 2016-05-11 1.78 Crystal structure of E.Coli DsbA in complex with 2-(4-iodophenylsulfonamido) benzoic acid
3DKS 2009-05-12 1.9 DsbA substrate complex
1ACV 1997-10-15 1.9 DSBA MUTANT H32S
2B3S 2006-09-05 1.96 structure of the DSBA mutant (P31G-C33A)
6BR4 2017-12-27 1.99 Crystal structure of Escherichia coli DsbA in complex with {N}-methyl-1-(3-thiophen-2-ylphenyl)methanamine
6BQX 2017-12-27 1.99 Crystal structure of Escherichia coli DsbA in complex with N-methyl-1-(4-phenoxyphenyl)methanamine
1U3A 2005-05-03 2.0 mutant DsbA
1DSB 1994-01-31 2.0 CRYSTAL STRUCTURE OF THE DSBA PROTEIN REQUIRED FOR DISULPHIDE BOND FORMATION IN VIVO
1A2J 1998-09-16 2.0 OXIDIZED DSBA CRYSTAL FORM II
1AC1 1997-10-15 2.0 DSBA MUTANT H32L
1FVJ 1997-05-15 2.06 THE 2.06 ANGSTROM STRUCTURE OF THE H32Y MUTANT OF THE DISULFIDE BOND FORMATION PROTEIN (DSBA)
1UN2 2003-09-26 2.4 Crystal structure of circularly permuted CPDSBA_Q100T99: Preserved Global Fold and Local Structural Adjustments
4TKY 2015-01-14 2.5 The complex structure of E. coli DsbA bound to a peptide at the DsbA/DsbB interface
1TI1 2005-05-03 2.6 crystal structure of a mutant DsbA
2B6M 2006-10-17 2.65 Structure of the DsbA mutant (P31A-C33A)
1A2L 1998-07-08 2.7 REDUCED DSBA AT 2.7 ANGSTROMS RESOLUTION
1A2M 1998-07-08 2.7 OXIDIZED DSBA AT 2.7 ANGSTROMS RESOLUTION, CRYSTAL FORM III
1BQ7 1999-08-20 2.8 DSBA MUTANT P151A, ROLE OF THE CIS-PROLINE IN THE ACTIVE SITE OF DSBA
2HI7 2006-12-05 3.7 Crystal structure of DsbA-DsbB-ubiquinone complex
3E9J 2008-11-25 3.7 Structure of the charge-transfer intermediate of the transmembrane redox catalyst DsbB
2ZUP 2009-04-14 3.7 Updated crystal structure of DsbB-DsbA complex from E. coli

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
99.5 Thiol:disulfide interchange protein DsbA P52235 DSBA_SHIFL
99.5 Thiol:disulfide interchange protein DsbA P0A4L5 DSBA_ECOL6
99.5 Thiol:disulfide interchange protein DsbA P0A4L6 DSBA_ECO27
100.0 Thiol:disulfide interchange protein DsbA P0AEG4 DSBA_ECOLI
100.0 Thiol:disulfide interchange protein DsbA P0AEG5 DSBA_ECO57