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Folding kinetics of a fluorescent variant of monomeric lambda repressor.

Publication ,  Journal Article
Ghaemmaghami, S; Word, JM; Burton, RE; Richardson, JS; Oas, TG
Published in: Biochemistry
June 23, 1998

A tryptophan-containing variant of monomeric lambda repressor has been made, and its folding kinetics were analyzed at 20 degreesC using fluorescence stopped-flow and dynamic NMR. Equilibrium denaturation curves obtained by circular dichroism, fluorescence, and NMR are superimposable. Stopped-flow analysis indicates that in the absence of denaturants the folding reaction is complete within the dead-time of the experiment. Within higher denaturant conditions, where the folding rate is slower, NMR and stopped-flow agree on the folding and unfolding rates of the protein. In 3.4 M urea and 1.8 M GdmCl, we show that the variant folds within 2 ms. Extrapolation indicates that the folding time is 20 micro(s) in the absence of denaturants. All folding and unfolding reactions displayed monoexponential kinetics, and no burst-phases were observed. In addition, the thermodynamic parameters Delta G and meq obtained from the kinetic analysis are consistent with the equilibrium experiments. The results support a two-state Dleft and right arrow N folding model.

Duke Scholars

Published In

Biochemistry

DOI

ISSN

0006-2960

Publication Date

June 23, 1998

Volume

37

Issue

25

Start / End Page

9179 / 9185

Location

United States

Related Subject Headings

  • Viral Regulatory and Accessory Proteins
  • Viral Proteins
  • Urea
  • Tryptophan
  • Spectrometry, Fluorescence
  • Repressor Proteins
  • Protein Folding
  • Protein Denaturation
  • Mutagenesis, Site-Directed
  • Models, Molecular
 

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Ghaemmaghami, S., Word, J. M., Burton, R. E., Richardson, J. S., & Oas, T. G. (1998). Folding kinetics of a fluorescent variant of monomeric lambda repressor. Biochemistry, 37(25), 9179–9185. https://doi.org/10.1021/bi980356b
Ghaemmaghami, S., J. M. Word, R. E. Burton, J. S. Richardson, and T. G. Oas. “Folding kinetics of a fluorescent variant of monomeric lambda repressor.Biochemistry 37, no. 25 (June 23, 1998): 9179–85. https://doi.org/10.1021/bi980356b.
Ghaemmaghami S, Word JM, Burton RE, Richardson JS, Oas TG. Folding kinetics of a fluorescent variant of monomeric lambda repressor. Biochemistry. 1998 Jun 23;37(25):9179–85.
Ghaemmaghami, S., et al. “Folding kinetics of a fluorescent variant of monomeric lambda repressor.Biochemistry, vol. 37, no. 25, June 1998, pp. 9179–85. Pubmed, doi:10.1021/bi980356b.
Ghaemmaghami S, Word JM, Burton RE, Richardson JS, Oas TG. Folding kinetics of a fluorescent variant of monomeric lambda repressor. Biochemistry. 1998 Jun 23;37(25):9179–9185.
Journal cover image

Published In

Biochemistry

DOI

ISSN

0006-2960

Publication Date

June 23, 1998

Volume

37

Issue

25

Start / End Page

9179 / 9185

Location

United States

Related Subject Headings

  • Viral Regulatory and Accessory Proteins
  • Viral Proteins
  • Urea
  • Tryptophan
  • Spectrometry, Fluorescence
  • Repressor Proteins
  • Protein Folding
  • Protein Denaturation
  • Mutagenesis, Site-Directed
  • Models, Molecular