In vitro folding of recombinant hepatitis B virus X-protein produced in Escherichia coli: formation of folding intermediates

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Title
In vitro folding of recombinant hepatitis B virus X-protein produced in Escherichia coli: formation of folding intermediates
Author(s)
Sun Ok Kim; Mi Jin Sohn; Soon Seog Jeong; Jeh Hoon Shin; Young Ik Lee
Bibliographic Citation
BMB Reports, vol. 32, no. 6, pp. 521-528
Publication Year
1999
Abstract
The folding of recombinant hepatitis B virus X-protein (rHBx) solubilized from Escherichia coli inclusion bodies was investigated. By sequential dialysis of urea, rHBx was folded into its native structure, which was demonstrated by the efficacy of its transcriptional activation of the adenovirus major late promoter (MLP), fluorescence spectroscopy, and circular dichroism (CD) analysis. The decrease in CD values at 220 nm and a corresponding blue shift of the intrinsic fluorescence emission confirmed the ability of rHBx to refold in lower concentrations of urea, yielding the active protein. Equilibrium and kinetic studies of the refolding of rHBx were carried out by tryptophan fluorescence measurements. From the biphasic nature of the fluorescence curves, the existence of stable intermediate states in the renaturation process was inferred. Reverse phase-high performance liquid chromatography (RP-HPLC) analysis further demonstrated the existence of these intermediates and their apparent compactness.
Keyword
Circular dichroismFluorescence emission maximumHepatitis B virus X-proteinRefoldingReverse phase-high performance liquid chromatography
ISSN
1225-8687
Publisher
Korea Soc-Assoc-Inst
Type
Article
Appears in Collections:
1. Journal Articles > Journal Articles
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