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Cloning, Functional Characterization and Site-Directed Mutagenesis of 4-Coumarate: Coenzyme A Ligase (4CL) Involved in Coumarin Biosynthesis in Peucedanum praeruptorum Dunn

Overview of attention for article published in Frontiers in Plant Science, January 2017
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Title
Cloning, Functional Characterization and Site-Directed Mutagenesis of 4-Coumarate: Coenzyme A Ligase (4CL) Involved in Coumarin Biosynthesis in Peucedanum praeruptorum Dunn
Published in
Frontiers in Plant Science, January 2017
DOI 10.3389/fpls.2017.00004
Pubmed ID
Authors

Tingting Liu, Ruolan Yao, Yucheng Zhao, Sheng Xu, Chuanlong Huang, Jun Luo, Lingyi Kong

Abstract

Coumarins are the main bioactive compounds in Peucedanum praeruptorum Dunn, a common Chinese herbal medicine. Nevertheless, the genes involved in the biosynthesis of core structure of coumarin in P. praeruptorum have not been identified yet. 4-Coumarate: CoA ligase (4CL) catalyzes the formation of hydroxycinnamates CoA esters, and plays an essential role at the divergence point from general phenylpropanoid metabolism to major branch pathway of coumarin. Here, three novel putative 4CL genes (Pp4CL1, Pp4CL7, and Pp4CL10) were isolated from P. praeruptorum. Biochemical characterization of the recombinant proteins revealed that Pp4CL1 utilized p-coumaric and ferulic acids as its two main substrates for coumarin biosynthesis in P. praeruptorum. Furthermore, Pp4CL1 also exhibited activity toward caffeic, cinnamic, isoferulic, and o-coumaric acids and represented a bona fide 4CL. Pp4CL7 and Pp4CL10 had no catalytic activity toward hydroxycinnamic acid compounds. But they had close phylogenetic relationship to true 4CLs and were defined as 4CL-like genes. Among all putative 4CLs, Pp4CL1 was the most highly expressed gene in roots, and its expression level was significantly up-regulated in mature roots compared with seedlings. Subcellular localization studies showed that Pp4CL1 and Pp4CL10 proteins were localized in the cytosol. In addition, site-directed mutagenesis of Pp4CL1 demonstrated that amino acids of Tyr-239, Ala-243, Met-306, Ala-309, Gly-334, Lys-441, Gln-446, and Lys-526 were essential for substrate binding or catalytic activities. The characterization and site-directed mutagenesis studies of Pp4CL1 lays a solid foundation for elucidating the biosynthetic mechanisms of coumarins in P. praeruptorum and provides further insights in understanding the structure-function relationships of this important family of proteins.

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Geographical breakdown

Country Count As %
Unknown 36 100%

Demographic breakdown

Readers by professional status Count As %
Student > Ph. D. Student 6 17%
Researcher 5 14%
Student > Bachelor 4 11%
Other 3 8%
Student > Master 3 8%
Other 4 11%
Unknown 11 31%
Readers by discipline Count As %
Biochemistry, Genetics and Molecular Biology 9 25%
Agricultural and Biological Sciences 8 22%
Chemistry 4 11%
Business, Management and Accounting 1 3%
Pharmacology, Toxicology and Pharmaceutical Science 1 3%
Other 1 3%
Unknown 12 33%
Attention Score in Context

Attention Score in Context

This research output has an Altmetric Attention Score of 1. This is our high-level measure of the quality and quantity of online attention that it has received. This Attention Score, as well as the ranking and number of research outputs shown below, was calculated when the research output was last mentioned on 11 February 2017.
All research outputs
#20,403,545
of 22,953,506 outputs
Outputs from Frontiers in Plant Science
#16,278
of 20,388 outputs
Outputs of similar age
#354,033
of 418,227 outputs
Outputs of similar age from Frontiers in Plant Science
#384
of 516 outputs
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