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Clinical and Diagnostic Laboratory Immunology, January 2000, p. 58-63, Vol. 7, No. 1
1071-412X/0/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Characterization of a Monoclonal Antibody and Its Single-Chain Antibody Fragment Recognizing the Nucleoside Triphosphatase/Helicase Domain of the Hepatitis C Virus Nonstructural 3 Protein

Zhu-Xu Zhang,1 Una Lazdina,1 Margaret Chen,1 Darrell L. Peterson,2 and Matti Sällberg1,*

Divisions of Clinical Virology and Basic Oral Sciences, Huddinge University Hospital, S-141 86 Huddinge, Sweden,1 and Department of Biochemistry and Molecular Biophysics, Virginia Commonwealth University, Richmond, Virginia2

Received 6 July 1999/Returned for modification 19 August 1999/Accepted 6 October 1999

We have produced a murine monoclonal antibody (MAb), ZX10, recognizing the NTPase/helicase domain of the hepatitis C virus (HCV) nonstructural 3 protein (NS3), from which we designed a single-chain variable fragment (ScFv). The ZX10 MAb recognized a discontinuous epitope of the NTPase/helicase domain, of which the linear sequence GEIPFYGKAIPL at residues 1371 to 1382 constitutes one part. cDNAs from variable regions coding for the heavy and light chains were cloned, sequenced, and assembled into the NS3-ScFv, which was inserted into procaryotic and eucaryotic expression vectors. Escherichia coli-expressed NS3-ScFv inhibited the binding of the ZX10 MAb to NS3, confirming a retained specificity. However, the ability to bind the peptide 1371-1382 had been lost. In vitro-translated NS3-ScFv and HCV NS3/NS4A were coprecipitated by antibodies to HCV NS4A, confirming the in vitro activity of the NS3 ScFv. Thus, we have designed a functional NS3 NTPase/helicase domain-specific ScFv which should be evaluated further with respect to disturbing enzymatic functions of the NS3 protein.

* Corresponding author. Mailing address: Division of Clinical Virology, F 68, Huddinge University Hospital, S-141 86 Huddinge, Sweden. Phone: 46-8-5858 7939. Fax: 46-8-5858 7933. E-mail: misg{at}labd01.hs.sll.se.

Clinical and Diagnostic Laboratory Immunology, January 2000, p. 58-63, Vol. 7, No. 1
1071-412X/0/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


This article has been cited by other articles:

  • Lazdina, U., Cao, T., Steinbergs, J., Alheim, M., Pumpens, P., Peterson, D. L., Milich, D. R., Leroux-Roels, G., Sallberg, M. (2001). Molecular Basis for the Interaction of the Hepatitis B Virus Core Antigen with the Surface Immunoglobulin Receptor on Naive B Cells. J. Virol. 75: 6367-6374 [Abstract] [Full Text]  
  • Lazdina, U., Hultgren, C., Frelin, L., Chen, M., Lodin, K., Weiland, O., Leroux-Roels, G., Quiroga, J. A., Peterson, D. L., Milich, D. R., Sällberg, M. (2001). Humoral and CD4+ T helper (Th) cell responses to the hepatitis C virus non-structural 3 (NS3) protein: NS3 primes Th1-like responses more effectively as a DNA-based immunogen than as a recombinant protein. J. Gen. Virol. 82: 1299-1308 [Abstract] [Full Text]  


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