High-throughput screening assay for d-amino acid oxidase

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• 作者：Svetlana V. Khoronenkova ; Vladimir I. Tishkov
• 关键词：d-Amino acid oxidase ; High-throughput screening ; Peroxidase reaction ; Membrane ; Recombinant Escherichia coli cells ; Mutant library
• 刊名：Analytical Biochemistry
• 出版年：2008
• 出版时间：15 March 2008
• 年：2008
• 卷：374
• 期：2
• 页码：405-410
• 全文大小：1062 K

文摘

A membrane-based high-throughput screening (HTS) assay for active d-amino acid oxidase (DAAO) in liquid samples as well as in intact Escherichia coli cells has been developed and optimized. The detection limit of the assay was less than 1 ng per sample. The method proposed can be used for quantitative DAAO determination in the range of 0.13 to 3.60 ng enzyme per probe. The protocol was successfully tested to screen a library of E. coli clones containing mutant DAAOs active toward target substrates.