Dynamic equilibrium unfolding pathway of human tumor necrosis factor-α induced by guanidine hydrochloride

Yong Rok Kim, Joong Sik Hahn, Heedeok Hong, Woojin Jeong, Nam Woong Song, Hang Cheol Shin, Dongho Kim

Research output: Contribution to journalArticle

6 Citations (Scopus)

Abstract

The dynamic equilibrium unfolding pathway of human tumor necrosis factor-α (TNF-α) during denaturation at different guanidine hydrochloride (GdnHCl) concentrations (0-4.2 M) was investigated by steady-state fluorescence spectroscopy, potassium iodide (KI) fluorescence quenching, far-UV circular dichroism (CD), picosecond time-resolved fluorescence lifetime, and anisotropy decay measurements. We utilized the intrinsic fluorescence of Trp-28 and Trp-114 to characterize the conformational changes involved in the equilibrium unfolding pathway. The detailed unfolding pathway under equilibrium conditions was discussed with respect to motional dynamics and partially folded structures. At 0-0.9 M [GdnHCl], the rotational correlation times of 22-25 ns were obtained from fluorescence anisotropy decay measurements and assigned to those of trimeric states by hydrodynamic calculation. In this range, the solvent accessibility of Trp residues increased with increasing [GdnHCl], suggesting the slight expansion of the trimeric structure. At 1.2-2.1 M [GdnHCl], the enhanced solvent accessibility and the rotational degree of freedom of Trp residues were observed, implying the loosening of the internal structure. In this [GdnHCl] region, TNF-α was thought to be in soluble aggregates having distinct conformational characteristics from a native (N) or fully unfolded state (U). At 4.2 M [GdnHCl], TNF-α unfolded to a U-state. From these results, the equilibrium unfolding pathway of TNF-α, trimeric and all β-sheet protein, could not be viewed from the simple two state model (N→U). Copyright (C) 1999 Elsevier Science B.V.

Original languageEnglish
Pages (from-to)486-495
Number of pages10
JournalBiochimica et Biophysica Acta - Protein Structure and Molecular Enzymology
Volume1429
Issue number2
DOIs
Publication statusPublished - 1999 Jan 11

Fingerprint

Guanidine
Fluorescence
Fluorescence Polarization
Tumor Necrosis Factor-alpha
Anisotropy
Potassium Iodide
Denaturation
Fluorescence Spectrometry
Fluorescence spectroscopy
Hydrodynamics
Circular Dichroism
human TNF protein
Quenching
Proteins

All Science Journal Classification (ASJC) codes

  • Structural Biology
  • Biophysics
  • Biochemistry
  • Molecular Biology

Cite this

@article{aa0375891c3a4e07b7e0dab0c5d52aef,
title = "Dynamic equilibrium unfolding pathway of human tumor necrosis factor-α induced by guanidine hydrochloride",
abstract = "The dynamic equilibrium unfolding pathway of human tumor necrosis factor-α (TNF-α) during denaturation at different guanidine hydrochloride (GdnHCl) concentrations (0-4.2 M) was investigated by steady-state fluorescence spectroscopy, potassium iodide (KI) fluorescence quenching, far-UV circular dichroism (CD), picosecond time-resolved fluorescence lifetime, and anisotropy decay measurements. We utilized the intrinsic fluorescence of Trp-28 and Trp-114 to characterize the conformational changes involved in the equilibrium unfolding pathway. The detailed unfolding pathway under equilibrium conditions was discussed with respect to motional dynamics and partially folded structures. At 0-0.9 M [GdnHCl], the rotational correlation times of 22-25 ns were obtained from fluorescence anisotropy decay measurements and assigned to those of trimeric states by hydrodynamic calculation. In this range, the solvent accessibility of Trp residues increased with increasing [GdnHCl], suggesting the slight expansion of the trimeric structure. At 1.2-2.1 M [GdnHCl], the enhanced solvent accessibility and the rotational degree of freedom of Trp residues were observed, implying the loosening of the internal structure. In this [GdnHCl] region, TNF-α was thought to be in soluble aggregates having distinct conformational characteristics from a native (N) or fully unfolded state (U). At 4.2 M [GdnHCl], TNF-α unfolded to a U-state. From these results, the equilibrium unfolding pathway of TNF-α, trimeric and all β-sheet protein, could not be viewed from the simple two state model (N→U). Copyright (C) 1999 Elsevier Science B.V.",
author = "Kim, {Yong Rok} and Hahn, {Joong Sik} and Heedeok Hong and Woojin Jeong and Song, {Nam Woong} and Shin, {Hang Cheol} and Dongho Kim",
year = "1999",
month = "1",
day = "11",
doi = "10.1016/S0167-4838(98)00263-5",
language = "English",
volume = "1429",
pages = "486--495",
journal = "Biochimica et Biophysica Acta - Proteins and Proteomics",
issn = "1570-9639",
publisher = "Elsevier",
number = "2",

}

Dynamic equilibrium unfolding pathway of human tumor necrosis factor-α induced by guanidine hydrochloride. / Kim, Yong Rok; Hahn, Joong Sik; Hong, Heedeok; Jeong, Woojin; Song, Nam Woong; Shin, Hang Cheol; Kim, Dongho.

In: Biochimica et Biophysica Acta - Protein Structure and Molecular Enzymology, Vol. 1429, No. 2, 11.01.1999, p. 486-495.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Dynamic equilibrium unfolding pathway of human tumor necrosis factor-α induced by guanidine hydrochloride

AU - Kim, Yong Rok

AU - Hahn, Joong Sik

AU - Hong, Heedeok

AU - Jeong, Woojin

AU - Song, Nam Woong

AU - Shin, Hang Cheol

AU - Kim, Dongho

PY - 1999/1/11

Y1 - 1999/1/11

N2 - The dynamic equilibrium unfolding pathway of human tumor necrosis factor-α (TNF-α) during denaturation at different guanidine hydrochloride (GdnHCl) concentrations (0-4.2 M) was investigated by steady-state fluorescence spectroscopy, potassium iodide (KI) fluorescence quenching, far-UV circular dichroism (CD), picosecond time-resolved fluorescence lifetime, and anisotropy decay measurements. We utilized the intrinsic fluorescence of Trp-28 and Trp-114 to characterize the conformational changes involved in the equilibrium unfolding pathway. The detailed unfolding pathway under equilibrium conditions was discussed with respect to motional dynamics and partially folded structures. At 0-0.9 M [GdnHCl], the rotational correlation times of 22-25 ns were obtained from fluorescence anisotropy decay measurements and assigned to those of trimeric states by hydrodynamic calculation. In this range, the solvent accessibility of Trp residues increased with increasing [GdnHCl], suggesting the slight expansion of the trimeric structure. At 1.2-2.1 M [GdnHCl], the enhanced solvent accessibility and the rotational degree of freedom of Trp residues were observed, implying the loosening of the internal structure. In this [GdnHCl] region, TNF-α was thought to be in soluble aggregates having distinct conformational characteristics from a native (N) or fully unfolded state (U). At 4.2 M [GdnHCl], TNF-α unfolded to a U-state. From these results, the equilibrium unfolding pathway of TNF-α, trimeric and all β-sheet protein, could not be viewed from the simple two state model (N→U). Copyright (C) 1999 Elsevier Science B.V.

AB - The dynamic equilibrium unfolding pathway of human tumor necrosis factor-α (TNF-α) during denaturation at different guanidine hydrochloride (GdnHCl) concentrations (0-4.2 M) was investigated by steady-state fluorescence spectroscopy, potassium iodide (KI) fluorescence quenching, far-UV circular dichroism (CD), picosecond time-resolved fluorescence lifetime, and anisotropy decay measurements. We utilized the intrinsic fluorescence of Trp-28 and Trp-114 to characterize the conformational changes involved in the equilibrium unfolding pathway. The detailed unfolding pathway under equilibrium conditions was discussed with respect to motional dynamics and partially folded structures. At 0-0.9 M [GdnHCl], the rotational correlation times of 22-25 ns were obtained from fluorescence anisotropy decay measurements and assigned to those of trimeric states by hydrodynamic calculation. In this range, the solvent accessibility of Trp residues increased with increasing [GdnHCl], suggesting the slight expansion of the trimeric structure. At 1.2-2.1 M [GdnHCl], the enhanced solvent accessibility and the rotational degree of freedom of Trp residues were observed, implying the loosening of the internal structure. In this [GdnHCl] region, TNF-α was thought to be in soluble aggregates having distinct conformational characteristics from a native (N) or fully unfolded state (U). At 4.2 M [GdnHCl], TNF-α unfolded to a U-state. From these results, the equilibrium unfolding pathway of TNF-α, trimeric and all β-sheet protein, could not be viewed from the simple two state model (N→U). Copyright (C) 1999 Elsevier Science B.V.

UR - http://www.scopus.com/inward/record.url?scp=0032942640&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0032942640&partnerID=8YFLogxK

U2 - 10.1016/S0167-4838(98)00263-5

DO - 10.1016/S0167-4838(98)00263-5

M3 - Article

VL - 1429

SP - 486

EP - 495

JO - Biochimica et Biophysica Acta - Proteins and Proteomics

JF - Biochimica et Biophysica Acta - Proteins and Proteomics

SN - 1570-9639

IS - 2

ER -