John E O'Connell

US Patent:

58916210, Apr 6, 1999

Filed:

Sep 24, 1997

Appl. No.:

8/936646

Inventors:

Renee M. Chabin - Neptune NJ
David W. Kuo - Princeton NJ
John F. O'Connell - Cranbury NJ
David L. Pompliano - Lawrenceville NJ
Kenny K. Wong - Edison NJ

Assignee:

Merck & Co., Inc. - Rahway NJ

International Classification:

C12Q 100
C12Q 137
C12Q 118
C12Q 148

US Classification:

435 4

Abstract:

An in vitro screening assay which identifies enzyme inhibitors and allows for the simultaneous assay of many enzymes. Enzyme, substrate, co-factor, etc. concentrations are optimized so that inhibitors of any one of the enzymes in the pathway are equally likely to be detected. Necessarily, the flux of substrate through each enzyme should be nearly the same during the assay, i. e. , each of the enzyme catalyzed steps must be equally rate-limiting. Preferably, optimal assay conditions are predicted by computer modeling. Further, the pathway conditions are optimized through variation of enzyme, starting substrate, co-substrate and co-factor concentrations. A positive response is initially detected as a change in the amount of the product generated at the end of the enzyme cascade as compared to a standard. A sample producing a positive result can be deconvoluted.