An enzyme-linked immunosorbent assay for detecting proteolytic activity of hepatitis C virus proteinase

Analytical Biochemistry
N TakeshitaK Shimotohno

Abstract

An ELISA method for the quantitation in vitro of HCV serine proteinase activity was developed. A peptide substrate, Ac-Gly-Glu-Ala-Gly-Asp-Asp-Ile-Val-Pro-Cys-Ser-Met-Ser-Tyr-Thr-Trp-Thr-L ys (biotin) -OH (Sub-1), was hydrolyzed by a recombinant NS3 proteinase fused with maltose binding protein (MBP-NS3) into a product with a free amino moiety at the N-terminus. The product was immobilized, and the amino moiety was analyzed by digoxigenin labeling followed by immunological reaction with anti-digoxigenin-alkaline phosphatase conjugate and then the colorimeteric reaction. This method is suited for the high throughput screening of inhibitors, and the screening can be accelerated by automatic operation.

References

Aug 1, 1991·Hepatology : Official Journal of the American Association for the Study of Liver Diseases·M HoughtonQ L Choo
Mar 15, 1991·Proceedings of the National Academy of Sciences of the United States of America·Q L ChooP J Barr
Dec 1, 1990·Proceedings of the National Academy of Sciences of the United States of America·N KatoK Shimotohno
Nov 15, 1993·Proceedings of the National Academy of Sciences of the United States of America·M HijikataK Shimotohno
Jul 26, 1995·Biochemical and Biophysical Research Communications·T MizutaniK Shimotohno
Aug 15, 1995·Proceedings of the National Academy of Sciences of the United States of America·C Lin, C M Rice
May 25, 1995·Biochemical and Biophysical Research Communications·N KakiuchiK Shimotohno

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Citations

Sep 27, 2005·Assay and Drug Development Technologies·Jin-Ching LeeJohn T-A Hsu
Jul 2, 2005·Expert Opinion on Emerging Drugs·B W Dymock
Jul 14, 2009·Analytical Biochemistry·Shawn MisialekKarl Kossen
Oct 17, 2002·Analytical Biochemistry·Barbara MouratouJean-Luc Guesdon
Feb 19, 1998·FEBS Letters·N KakiuchiK Shimotohno
Jan 16, 1999·Analytical Biochemistry·M CerretaniS Altamura
May 21, 1999·Analytical Biochemistry·R ZhangB A Malcolm
Jul 14, 1999·Journal of Virological Methods·N KakiuchiK Shimotohno
Dec 24, 1998·Antiviral Research·A D KwongS A Raybuck

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