Folding and stability of sweet protein single-chain monellin: An insight to protein engineering

Yoon Hui Sung, Hee Deok Hong, Chaejoon Cheong, Jin Hwan Kim, Joong Myung Cho, Yong Rok Kim, Weon Tae Lee

Research output: Contribution to journalArticle

14 Citations (Scopus)

Abstract

Engineered single-chain monellin (SCM) proteins were constructed by recombinant technology without disrupting the topology and sweet activity of native protein. Data from 8-anilinonaphthalene-1-sulfonic acid fluorescence, size-exclusion chromatography, and heteronuclear NMR strongly suggest the presence of a folding intermediate at 1.5 M GdnHCl for SCM protein. The structural feature of the folding intermediate from NMR data reveals that the secondary structures became mostly unstable, and protein experiences a dynamic equilibrium between native and unfolded state. All backbone amide protons exchange within 10 min, which imply that no stable hydrogen bonds exist in the secondary structural regions in the folding intermediate. From equilibrium unfolding and mutagenesis studies, the unfolding transition midpoints of mutant proteins gradually shifted toward lower denaturant concentration, indicating stability reductions of mutant proteins. Our results suggest that stability and folding pathways of SCM proteins could be regulated by a combined study of spectroscopy and mutagenesis, and these studies will provide useful information for understanding the folding kinetics of novel engineered proteins.

Original languageEnglish
Pages (from-to)44229-44238
Number of pages10
JournalJournal of Biological Chemistry
Volume276
Issue number47
DOIs
Publication statusPublished - 2001 Nov 23

Fingerprint

Protein Engineering
Protein Stability
Mutagenesis
Proteins
Mutant Proteins
Nuclear magnetic resonance
Biomolecular Nuclear Magnetic Resonance
Size exclusion chromatography
Amides
Gel Chromatography
Protons
Hydrogen
Spectrum Analysis
Hydrogen bonds
Fluorescence
Topology
Spectroscopy
Technology
Kinetics

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

Cite this

Sung, Yoon Hui ; Hong, Hee Deok ; Cheong, Chaejoon ; Kim, Jin Hwan ; Cho, Joong Myung ; Kim, Yong Rok ; Lee, Weon Tae. / Folding and stability of sweet protein single-chain monellin : An insight to protein engineering. In: Journal of Biological Chemistry. 2001 ; Vol. 276, No. 47. pp. 44229-44238.
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Folding and stability of sweet protein single-chain monellin : An insight to protein engineering. / Sung, Yoon Hui; Hong, Hee Deok; Cheong, Chaejoon; Kim, Jin Hwan; Cho, Joong Myung; Kim, Yong Rok; Lee, Weon Tae.

In: Journal of Biological Chemistry, Vol. 276, No. 47, 23.11.2001, p. 44229-44238.

Research output: Contribution to journalArticle

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T2 - An insight to protein engineering

AU - Sung, Yoon Hui

AU - Hong, Hee Deok

AU - Cheong, Chaejoon

AU - Kim, Jin Hwan

AU - Cho, Joong Myung

AU - Kim, Yong Rok

AU - Lee, Weon Tae

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