Advanced Search

Journal Navigation

Journal Home

Subscriptions

Archive

Contact Us

Table of Contents

Click here for more information

CiteULike is a free service for managing and discovering scholarly references - click here to get started.

Sign In to gain access to subscriptions and/or personal tools.
Journal of Biomolecular Screening
This Article
Right arrow Full Text (PDF)
Right arrow References
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Add to Saved Citations
Right arrow Download to citation manager
Right arrowRequest Permissions
Right arrow Request Reprints
Right arrow Add to My Marked Citations
Citing Articles
Right arrow Citing Articles via Web of Science (4)
Right arrow Citing Articles via Google Scholar
Right arrow Citing Articles via Scopus
Google Scholar
Right arrow Articles by Fowler, A.
Right arrow Articles by Schulman, M.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Fowler, A.
Right arrow Articles by Schulman, M.
Social Bookmarking
Add to CiteULike   Add to Complore   Add to Connotea   Add to Del.icio.us   Add to Digg   Add to Reddit   Add to Technorati   Add to Twitter  
What's this?

Development of a High Throughput Scintillation Proximity Assay for Hepatitis C Virus NS3 Protease That Reduces the Proportion of Competitive Inhibitors Identified

Anne Fowler

Department of SPA Technology, Amersham Pharmacia Biotech UK Limited, Little Chalfont, Buckinghamshire, England

Molly Price-Jones

Department of SPA Technology, Amersham Pharmacia Biotech UK Limited, Little Chalfont, Buckinghamshire, England

Kelvin Hughes

Department of SPA Technology, Amersham Pharmacia Biotech UK Limited, Little Chalfont, Buckinghamshire, England

John Anson

Department of SPA Technology, Amersham Pharmacia Biotech UK Limited, Little Chalfont, Buckinghamshire, England

Russell Lingham

Merck Research Laboratories, Rahway, NJ

Marvin Schulman

Merck Research Laboratories, Rahway, NJ

A screening assay has been developed for hepatitis C virus (HCV) NS3 protease using the scintillation proximity assay (SPA) technology. The sequence of the peptide substrate used was taken from the site cleaved by the enzyme in the mature nonstructural protein of HCV. The peptide was biotinylated at the N-terminus and tritiated at the C-terminus so that a decrease in signal was detected as a result of enzyme activity.

IC50 values were calculated for the cleaved product, and it was shown that the value obtained was dependent on the substrate concentration used. The effect of substrate concentration on the inhibition of HCV NS3 protease was further highlighted in a mock screening assay, using colored natural product samples, in which the hit rate was altered by a change in substrate concentration. An increase in substrate concentration reduced the proportion of competitive inhibitors identified. This study highlighted the importance of optimizing the components used in SPA assays in order to obtain an assay format valid for high throughput screening.

Journal of Biomolecular Screening, Vol. 5, No. 3, 153-158 (2000)
DOI: 10.1177/108705710000500307
Add to CiteULike CiteULike   Add to Complore Complore   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us   Add to Digg Digg   Add to Reddit Reddit   Add to Technorati Technorati   Add to Twitter Twitter    What's this?