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- Title
An in vitro screening assay to discover novel inhibitors of 4-coumarate:CoA ligase.
- Authors
Min-Soo Yun; Weijun Chen; Fan Deng; Kiyokawa, Takahiro; Mametsuka, Koki; Yogo, Yasuhiro
- Abstract
4-Coumarate:CoA ligase (4CL, EC 6.2.1.12) exists only in plants and plays an important role in the phenylpropanoid pathway. Identification of inhibitors targeting 4CL provides a novel approach for developing effective plant growth inhibitors (PGIs). The full-length gene of tobacco 4CL (Nt4CL1) was cloned and expressed in Escherichia coli Cast & Chalm. The recombinant 4CL protein was extracted and purified by several purification steps including gel-filtration and anion-exchange chromatography. 4CL activity assay was miniaturized and optimized using a 96-well microplate and a reader. Among 28 existing herbicides, propanil and swep strongly inhibited in vitro 4CL enzyme activity, and they were selected for further studies. The process of this assay can be developed into a high-throughput screening system of PGI targeting 4CL in the phenyipropanoid pathway.
- Subjects
LIGASES; ENZYMES; BIOLOGICAL assay; PLANT growth; PLANT regulators; TOBACCO; ESCHERICHIA coli; CHROMATOGRAPHIC analysis; MICROPLATES; HERBICIDES
- Publication
Pest Management Science, 2006, Vol 62, Issue 11, p1065
- ISSN
1526-498X
- Publication type
Article
- DOI
10.1002/ps.1277