Structure and stability of an early folding intermediate of Escherichia coli trp aporepressor measured by far-UV stopped-flow circular dichroism and 8-anilino-1-naphthalene sulfonate binding.


Abstract

The refolding kinetics of Escherichia coli trp aporepressor were monitored using stopped-flow far-ultraviolet circular dichroism and 8-anilino-1-naphthalene sulfonate fluorescence spectroscopy. Significant gains in secondary structure and the development of hydrophobic surface, respectively, were observed within the dead time of mixing (4-5 ms). These initial increases, or burst phase amplitudes, plotted as a function of final urea concentration, exhibited sigmoidal, coincident unfolding transition curves. The transition curves were fit to a two-state model, and the resulting free energies of folding in the absence of denaturant were found to be similar (approximately 3.3 kcal/mol). Three subsequent slow refolding phases exhibited relaxation times and amplitudes similar to those previously observed for tryptophan fluorescence [Gittelman, M. S., & Matthews, C. R. (1990) Biochemistry 29, 7011-7021]. These results support the proposals that a stable, monomeric intermediate is rapidly formed during the folding of trp aporepressor and that this species contains a significant amount of secondary structure and hydrophobic surface. This early intermediate is then processed through folding and association reactions that result in the formation of the remaining secondary, tertiary, and quaternary structure. Study holds ProTherm entries: 4574 Extra Details: additive : Na2EDTA(0.1 mM), secondary structure; hydrophobic surface; two-state model;,association reactions

Submission Details

ID: 3AVQmUEb

Submitter: Connie Wang

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

Version: 1

Publication Details
Mann CJ;Matthews CR,Biochemistry (1993) Structure and stability of an early folding intermediate of Escherichia coli trp aporepressor measured by far-UV stopped-flow circular dichroism and 8-anilino-1-naphthalene sulfonate binding. PMID:8499433
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
6EJZ 2017-09-24T00:00:00+0000 1.9 Tryptophan Repressor TrpR from E.coli variant S88Y with Indole-3-acetic acid as ligand
2XDI 2010-05-03T00:00:00+0000 0 Tryptophan repressor with L75F mutation in its apo form (no L- tryptophan bound)
5TM0 2016-10-12T00:00:00+0000 0 Solution NMR structures of two alternative conformations of E. coli tryptophan repressor in dynamic equilibrium
1CO0 1999-05-30T00:00:00+0000 0 NMR STUDY OF TRP REPRESSOR-MTR OPERATOR DNA COMPLEX
1JHG 1996-07-19T00:00:00+0000 1.3 TRP REPRESSOR MUTANT V58I
1MI7 2002-08-22T00:00:00+0000 2.5 Crystal Structure of Domain Swapped trp Aporepressor in 30%(v/v) Isopropanol
1RCS 1995-05-12T00:00:00+0000 0 NMR STUDY OF TRP REPRESSOR-OPERATOR DNA COMPLEX
1TRO 1992-08-30T00:00:00+0000 1.9 CRYSTAL STRUCTURE OF TRP REPRESSOR OPERATOR COMPLEX AT ATOMIC RESOLUTION
1TRR 1993-10-21T00:00:00+0000 2.4 TANDEM BINDING IN CRYSTALS OF A TRP REPRESSOR/OPERATOR HALF-SITE COMPLEX
1WRP 1987-12-01T00:00:00+0000 2.2 FLEXIBILITY OF THE DNA-BINDING DOMAINS OF TRP REPRESSOR

Relevant UniProtKB Entries

Percent Identity Matching Chains Protein Accession Entry Name
99.1 Trp operon repressor B1LEK0 TRPR_ECOSM
100.0 Trp operon repressor B7LNT5 TRPR_ESCF3
100.0 Trp operon repressor B7NH68 TRPR_ECOLU
100.0 Trp operon repressor B7NW74 TRPR_ECO7I
99.1 Trp operon repressor Q1R248 TRPR_ECOUT
99.1 Trp operon repressor Q8FA42 TRPR_ECOL6
99.1 Trp operon repressor Q0T8R8 TRPR_ECOL5
99.1 Trp operon repressor A1AJW2 TRPR_ECOK1
99.1 Trp operon repressor B7N2W3 TRPR_ECO81
99.1 Trp operon repressor B7MNK2 TRPR_ECO45
99.1 Trp operon repressor B7UR23 TRPR_ECO27
100.0 Trp operon repressor Q3YU01 TRPR_SHISS
100.0 Trp operon repressor P0A883 TRPR_SHIFL
100.0 Trp operon repressor Q0SX19 TRPR_SHIF8
100.0 Trp operon repressor Q327K2 TRPR_SHIDS
100.0 Trp operon repressor Q31SU5 TRPR_SHIBS
100.0 Trp operon repressor B2TZS6 TRPR_SHIB3
100.0 Trp operon repressor B6I6P1 TRPR_ECOSE
100.0 Trp operon repressor P0A881 TRPR_ECOLI
100.0 Trp operon repressor B1IS26 TRPR_ECOLC
100.0 Trp operon repressor A8A8C2 TRPR_ECOHS
100.0 Trp operon repressor B1XFK3 TRPR_ECODH
100.0 Trp operon repressor C4ZT75 TRPR_ECOBW
100.0 Trp operon repressor B7LXV6 TRPR_ECO8A
100.0 Trp operon repressor B5Z4S5 TRPR_ECO5E
100.0 Trp operon repressor P0A882 TRPR_ECO57
100.0 Trp operon repressor B7LEN9 TRPR_ECO55
100.0 Trp operon repressor A7ZVT5 TRPR_ECO24
90.4 Trp operon repressor A9MR96 TRPR_SALAR