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2008 | 55 | 1 | 57-66
Article title

Circular dichroism analysis for multidomain proteins: studies of the irreversible unfolding of Hepatitis C virus helicase

Content
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EN
Abstracts
EN
The non-structural protein 3 (NS3) of Hepatitis C virus (HCV) is a bifunctional enzyme with RNA-dependent NTPase/RNA helicase and serine protease activities, and thus represents a promising target for anti-HCV therapy. These functions are performed by two distinct moieties; the N-terminal protease domain and the C-terminal helicase domain that further folds into three structural subdomains. To obtain lower molecular mass proteins suitable for nuclear magnetic resonance studies of helicase-inhibitor complexes, helicase domains 1, 2, and 1+2 devoid of a hydrophobic β-loop were overexpressed and purified. Circular dichroism studies were carried out to confirm the secondary structure content and to determine thermodynamic parameters describing the stability of the proteins. Both thermal and GuHCl-induced unfolding experiments confirmed the multidomain organization of the helicase. The unfolding transition observed for domain 1+2 was in agreement with the model of two well-resolved successive steps corresponding to the independent unfolding of domains 1 and 2, respectively. In the case of the full-length helicase, the presence of domain 3 remarkably changed the transition profile, leading to fast and irreversible transformation of partially unfolded protein.
Publisher

Year
Volume
55
Issue
1
Pages
57-66
Physical description
Dates
published
2008
received
2007-09-22
revised
2007-12-13
accepted
2007-12-31
(unknown)
2008-01-23
Contributors
  • Institute of Biochemistry and Biophysics PAS, Warszawa, Poland
  • Institute of Biochemistry and Biophysics PAS, Warszawa, Poland
  • Institute of Biochemistry and Biophysics PAS, Warszawa, Poland
  • Institute of Biochemistry and Biophysics PAS, Warszawa, Poland
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Document Type
Publication order reference
Identifiers
YADDA identifier
bwmeta1.element.bwnjournal-article-abpv55p57kz
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