title: A high-throughput screening method for amino acid dehydrogenase.
abstract: A simple and rapid screening method for amino acid dehydrogenase (e.g., leucine dehydrogenase, LDH) has been developed. It relies on a competitive relationship between a non-fluorescent Cu(II)-calcein complex and amino acid (e.g., l-2-aminobutyric acid, l-ABA). When ABA was introduced to a Cu(II)-calcein solution, it bound with the Cu(II) ions and this released calcein from the complex, which was detected as strong fluorescence. The principle of this high-throughput screening method was validated by screening an LDH mutant library. Compared with other methods, this method provided much quicker l-ABA detection and screening for leucine dehydrogenase mutations.
authors.affiliation: Institute of Bioengineering, Zhejiang University of Technology, Hangzhou 310014, People's Republic of China; Engineering Research Center of Bioconversion and Biopurification of Ministry of Education, Zhejiang University of Technology, Hangzhou 310014, People's Republic of China.
authors.forename: Jian-Miao
authors.initials: JM
authors.lastname: Xu
authors.affiliation: Institute of Bioengineering, Zhejiang University of Technology, Hangzhou 310014, People's Republic of China; Engineering Research Center of Bioconversion and Biopurification of Ministry of Education, Zhejiang University of Technology, Hangzhou 310014, People's Republic of China.
authors.forename: Fang-Tian
authors.initials: FT
authors.lastname: Fu
authors.affiliation: Institute of Bioengineering, Zhejiang University of Technology, Hangzhou 310014, People's Republic of China; Engineering Research Center of Bioconversion and Biopurification of Ministry of Education, Zhejiang University of Technology, Hangzhou 310014, People's Republic of China.
authors.forename: Hai-Feng
authors.initials: HF
authors.lastname: Hu
authors.affiliation: Institute of Bioengineering, Zhejiang University of Technology, Hangzhou 310014, People's Republic of China; Engineering Research Center of Bioconversion and Biopurification of Ministry of Education, Zhejiang University of Technology, Hangzhou 310014, People's Republic of China. Electronic address: zhengyg@zjut.edu.cn.
authors.forename: Yu-Guo
authors.initials: YG
authors.lastname: Zheng
mesh_terms: D000613:Aminobutyrates / Q000378:metabolism; D056831:Coordination Complexes / Q000737:chemistry; D003300:Copper / Q000737:chemistry; D057075:Enzyme Assays / Q000379:methods*; D005452:Fluoresceins / Q000737:chemistry; D057166:High-Throughput Screening Assays; D007477:Ions / Q000737:chemistry; D050803:Leucine Dehydrogenase / Q000378:metabolism*; D013050:Spectrometry, Fluorescence
chemical_list: D000613:Aminobutyrates; D056831:Coordination Complexes; D005452:Fluoresceins; D007477:Ions; D003300:Copper; D050803:Leucine Dehydrogenase; C007740:fluorexon
keywords: Calcein; Cu(II); Leucine dehydrogenase; l-2-aminobutyric acid