30802-00-7Relevant articles and documents
Effect of flexible linker length on the activity of fusion protein 4-coumaroyl-CoA ligase::stilbene synthase
Guo, Huili,Yang, Yadong,Xue, Feiyan,Zhang, Hong,Huang, Tiran,Liu, Wenbin,Liu, Huan,Zhang, Fenqiang,Yang, Mingfeng,Liu, Chunmei,Lu, Heshu,Zhang, Yansheng,Ma, Lanqing
, p. 598 - 606 (2017)
In order to elucidate the effect of flexible linker length on the catalytic efficiency of fusion proteins, two short flexible peptide linkers of various lengths were fused between Arabidopsis thaliana 4-coumaroyl-CoA ligase (4CL) and Polygonum cuspidatum stilbene synthase (STS) to generate fusion proteins 4CL-(GSG)n-STS (n ≤ 5) and 4CL-(GGGGS)n-STS (n ≤ 4). The fusion proteins were expressed in both Escherichia coli and Saccharomyces cerevisiae, and their bioactivities were tested in vitro and in vivo using purified proteins and engineered strains, respectively. The catalytic efficiency of the fusions decreased gradually with the increase of GSG or GGGGS repeats. In both engineered S. cerevisiae and E. coli in vivo experiments, the capacity of resveratrol production decreased gradually with increasing linker length. In silico analysis showed that the prediction of homology models of fusion proteins was consistent with the in vitro and in vivo results.
Facile enzymic synthesis of caffeoyl CoA
Meng, Huabin,Campbell, Wilbur H.
, p. 605 - 608 (1997)
p-Hydroxycinnamic acid:CoA ligase (EC 6.2.1.12) was extracted from aspen xylem and its activity with several cinnamic acids was determined. The highest activity was found with p-hydroxycinnamic acid and the extract had reasonable activity with ferulic, ca
4-Coumarate:coenzyme A ligase isoform 3 from Piper nigrum (Pn4CL3) catalyzes the CoA thioester formation of 3,4-methylenedioxycinnamic and piperic acids
Jin, Zhehao,Wungsintaweekul, Juraithip,Kim, Sang-Hoon,Kim, Jeong-Han,Shin, Yongho,Ro, Dae-Kyun,Kim, Soo-Un
, p. 61 - 74 (2020/02/28)
Black pepper, dried green fruit of Piper nigrum L., is a household spice most popular in the world. Piperine, the pungency compound of black pepper, is proposed to partially arise from phenylpropanoid pathway. In the biosynthesis of piperine, 4-coumarate:CoA ligase (4CLs) must play a pivotal role in activating intermediate acids to corresponding CoA thioesters to serve as substrates. Based on transcriptome data, we isolated three P. nigrum 4CL isoforms (Pn4CL1, -2, and -3) from unripe peppercorn. These Pn4CLs were expressed in E. coli for in vitro enzyme assay with putative substrates, namely cinnamic, coumaric, ferulic, piperonylic, 3,4-methylenedioxycinnamic (3,4-MDCA), and piperic acids. Phylogenetic analysis and substrate usage study indicated that Pn4CL1, active towards coumaric and ferulic acids, belongs to class I 4CL for lignin synthesis. Pn4CL2 was a typical cinnamate-specific coumarate:CoA ligase-like (CLL) protein. The Pn4CL3, as class II enzyme, exhibited general 4CL activity towards coumaric and ferulic acids. However, Pn4CL3 was also active towards piperonylic acid, 3,4-MDCA, and piperic acid. Pn4CL3 possessed ～2.6 times higher catalytic efficiency (kcat/KM) towards 3,4-MDCA and piperic acid than towards coumaric and ferulic acids, suggesting its specific role in piperine biosynthesis. Different substrate preference among the Pn4CL isoforms can be explained by 3-dimensional protein structure modeling, which demonstrated natural variants in amino acid residues of binding pocket to accommodate different substrates. Quantitative PCR analysis of these isoforms indicated that Pn4CL1 transcript level was highest in the roots whereas Pn4CL2 in the fruits and Pn4CL3 in the leaves.
Coenzyme A-Conjugated Cinnamic Acids – Enzymatic Synthesis of a CoA-Ester Library and Application in Biocatalytic Cascades to Vanillin Derivatives
Dippe, Martin,Bauer, Anne-Katrin,Porzel, Andrea,Funke, Evelyn,Müller, Anna O.,Schmidt, Jürgen,Beier, Maria,Wessjohann, Ludger A.
supporting information, p. 5346 - 5350 (2019/11/29)
We present a bioorthogonal method for the ligation of coenzyme A (CoA) with cinnamic acids. The reaction, which is the initial step in the biosynthesis of a multitude of bioactive secondary metabolites, is catalyzed by a promiscuous plant ligase and yields CoA conjugates with different functionalization in high purity and without formation of by-products. Its applicability in biosynthetic cascades is shown for the direct transformation of cinnamic acids into natural benzaldehydes (like vanillin) or artificial derivatives (e. g. ethylvanillin). (Figure presented.).
