High-Throughput NMR-Based Screening with Competition Binding Experiments
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- 作者：Claudio Dalvit ; Maria Flocco ; Stefan Knapp ; Marina Mostardini ; Rita Perego ; Brian J. Stockman ; Marina Veronesi ; and Mario Varasi
- 刊名：Journal of the American Chemical Society
- 出版年：2002
- 出版时间：July 3, 2002
- 年：2002
- 卷：124
- 期：26
- 页码：7702 - 7709
- 全文大小：112K
- 年卷期：v.124,no.26(July 3, 2002)
- ISSN：1520-5126
文摘
The Achille's heel of ligand-based NMR screening methods is their failure to detect high-affinityligands and molecules that bind covalently to the receptor. We have developed a novel approach forperforming high-throughput screening with NMR spectroscopy that overcomes this limitation. The methodalso permits detection of potential high-affinity molecules that are only marginally soluble, thus significantlyenlarging the diversity of compounds amenable to NMR screening. The techniques developed utilizetransverse and/or selective longitudinal relaxation parameters in combination with competition bindingexperiments. Mathematical expressions are derived for proper setup of the NMR experiments and forextracting an approximate value of the binding constant for the identified ligand from a single-pointmeasurement. With this approach it is possible to screen thousands of compounds in a short period oftime against protein or DNA and RNA fragments. The methodology can also be applied for screening plantand fungi extracts.