A high-throughput screen utilizing the fluorescence of riboflavin for identification of lumazine synthase inhibitors

Jinhua Chen; Boris Illarionov; Adelbert Bacher; Markus Fischer; Ilka Haase; Gunda Georg; Qi Zhuang Ye; Zeqiang Ma; Mark Cushman

doi:10.1016/j.ab.2004.11.033

A high-throughput screen utilizing the fluorescence of riboflavin for identification of lumazine synthase inhibitors

Jinhua Chen, Boris Illarionov, Adelbert Bacher, Markus Fischer, Ilka Haase, Gunda Georg, Qi Zhuang Ye, Zeqiang Ma, Mark Cushman

Medicinal Chemistry

Research output: Contribution to journal › Article › peer-review

23 Scopus citations

Abstract

A high-throughput screening method based on the competitive binding of a lumazine synthase inhibitor and riboflavin to the active site of Schizosaccharomyces pombe lumazine synthase was developed. This assay is sensitive, simple, and robust. During assay development, all of the known active inhibitors tested were positively identified. Preliminary high-throughput screening in 384-well format resulted in a Z factor of 0.7. The approach utilizes a thermodynamic assay to bypass the problems associated with the instabilities of both lumazine synthase substrates that complicate the use of a kinetic assay in a high-throughput format, and it removes the time element from the assay, thus simplifying the procedure.

Original language	English (US)
Pages (from-to)	124-130
Number of pages	7
Journal	Analytical Biochemistry
Volume	338
Issue number	1
DOIs	https://doi.org/10.1016/j.ab.2004.11.033
State	Published - Mar 1 2005

Bibliographical note

Funding Information:
This research was made possible by NIH Grant GM51469 and by support from the Deutsche Forschungsgemeinschaft, Fonds der Chemischen Industrie, and the Hans Fischer Gesellschaft. The KU-High Throughput Screening Laboratory was established through funding from the National Institutes of Health COBRE award 1 P20 RR15563, from the State of Kansas, and the University of Kansas.

Keywords

Enzyme inhibitor discovery
High-throughput screening
Lumazine synthase
Riboflavin synthase

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abstract = "A high-throughput screening method based on the competitive binding of a lumazine synthase inhibitor and riboflavin to the active site of Schizosaccharomyces pombe lumazine synthase was developed. This assay is sensitive, simple, and robust. During assay development, all of the known active inhibitors tested were positively identified. Preliminary high-throughput screening in 384-well format resulted in a Z factor of 0.7. The approach utilizes a thermodynamic assay to bypass the problems associated with the instabilities of both lumazine synthase substrates that complicate the use of a kinetic assay in a high-throughput format, and it removes the time element from the assay, thus simplifying the procedure.",

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AU - Chen, Jinhua

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AU - Bacher, Adelbert

AU - Fischer, Markus

AU - Haase, Ilka

AU - Georg, Gunda

AU - Ye, Qi Zhuang

AU - Ma, Zeqiang

AU - Cushman, Mark

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AB - A high-throughput screening method based on the competitive binding of a lumazine synthase inhibitor and riboflavin to the active site of Schizosaccharomyces pombe lumazine synthase was developed. This assay is sensitive, simple, and robust. During assay development, all of the known active inhibitors tested were positively identified. Preliminary high-throughput screening in 384-well format resulted in a Z factor of 0.7. The approach utilizes a thermodynamic assay to bypass the problems associated with the instabilities of both lumazine synthase substrates that complicate the use of a kinetic assay in a high-throughput format, and it removes the time element from the assay, thus simplifying the procedure.

KW - Enzyme inhibitor discovery

KW - High-throughput screening

KW - Lumazine synthase

KW - Riboflavin synthase

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A high-throughput screen utilizing the fluorescence of riboflavin for identification of lumazine synthase inhibitors

Abstract

Bibliographical note

Keywords

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