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
Autor: | Matthews Cr, Mann Cj |
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Rok vydání: | 1993 |
Předmět: |
Circular dichroism
Protein Folding Kinetics Analytical chemistry Biochemistry Fluorescence spectroscopy Anilino Naphthalenesulfonates Protein Structure Secondary chemistry.chemical_compound Bacterial Proteins Drug Stability Escherichia coli Protein secondary structure Fluorescent Dyes Molecular Structure Circular Dichroism Escherichia coli Proteins Burst phase Hydrogen-Ion Concentration Folding (chemistry) Repressor Proteins Crystallography Sulfonate chemistry Thermodynamics Protein quaternary structure Apoproteins |
Zdroj: | Biochemistry. 32(20) |
ISSN: | 0006-2960 |
Popis: | 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. |
Databáze: | OpenAIRE |
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