Ligands for phosphatase binding assay
US6348572B1 · kind B1 · utility
Assignee
Inventors
Key dates
| Filing date | Apr 29, 1998 |
| Grant date | Feb 19, 2002 |
| Priority date | — |
| Expiry date | Apr 29, 2018 |
Classification
- Technology area (CPC G)Physics
- CPC primaryG11B7/25
- WIPO fieldAudio-visual technology
- WIPO sectorElectrical engineering
Abstract
Disclosed are new ligands for use in a binding assay for proteases and phosphatases, which contain cysteine in their binding sites or as a necessary structural component for enzymatic binding. The sulfhydryl group of cysteine is the nucleophilic group in the enzyme's mechanistic proteolytic and hydrolytic properties. The assay can be used to determine the ability of new, unknown ligands and mixtures of compounds to competitively bind with the enzyme versus a known binding agent for the enzyme, e.g., a known enzyme inhibitor. By the use of a mutant form of the natural or native wild-type enzyme, in which serine, or another amino acid, e.g., alanine, replaces cysteine, the problem of interference from extraneous oxidizing and alkylating agents in the assay procedure is overcome. The interference arises because of oxidation or alkylation of the sulfhydryl, —SH (or —S−), in the cysteine, which then adversely affects the binding ability of the enzyme. Specifically disclosed is an assay for tyrosine phosphatases and cysteine proteases, including caspases and cathepsins, e.g., Cathepsin K(O2), utilizing scintillation proximity assay (SPA) technology. The assay has import…
Source: USPTO / EPO open patent data. Objective bibliographic and citation counts.