Molecular cloning and biochemical characterization of a new coumarin glycosyltransferase CtUGT1 from Cistanche tubulosa

Xu, Xiping; Yan, Yaru; Huang, Wenqian; Mo, Ting; Wang, Xiaohui; Wang, Juan; Li, Jun; Shi, She‐Po; Liu, Xiao; Tu, Pengfei

doi:10.1016/j.fitote.2021.104995

Cited by 3 publications

(2 citation statements)

References 37 publications

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“…The function of MaBGLU1 was verified by overexpression in Melilotus , heterologous expression in E. coli and yeast two-hybrid experiments. Xu et al. (2021) cloned a new UGT from Cistanche tubulosa, and extensive in vitro enzyme assays found that CtUGTi could catalyze the glucosylation of coumarins umbelliferone 1, esculetine 2, and hymecromone 3.…”

Section: Survey Methodologymentioning

confidence: 99%

Anabolic metabolism of autotoxic substance coumarins in plants

Wu,

Shi,

Zhang

et al. 2023

PeerJ

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Background Autotoxicity is an intraspecific manifestation of allelopathy in plant species. The specialized metabolites and their derivatives that cause intraspecific allelopathic inhibition in the plant are known as autotoxic substances. Consequently, autotoxic substances production seriously affects the renewal and stability of ecological communities. Methods This article systematically summarizes the types of autotoxic substances present in different plants. They mainly include phenolic compounds, terpenoids, and nitrogenous organic compounds. Phenolic coumarins are the main autotoxic substances in many plants. Therefore, we also discuss differences in coumarin types and content among plant varieties, developmental stages, and tissue parts, as well as their mechanisms of autotoxicity. In addition, we review the metabolic pathways involved in coumarin biosynthesis, the key enzymes, genes, and transcription factors, as well as factors affecting coumarin biosynthesis. Results Coumarin biosynthesis involves three stages: (1) the formation of the coumarin nucleus; (2) acylation, hydroxylation, and cyclization; (3) structural modification. The key enzymes involved in the coumarin nuclear formation stage include PAL, C4H, 4CL, HCT, CAOMT, COSY, F6’H, and CCoAOMT1, and the key genes involved include BGA, CYP450 and MDR, among others. Ortho-hydroxylation is a key step in coumarin biosynthesis and PS, COSY and S8H are the key enzymes involved in this process. Finally, UGTs are responsible for the glycosylation modification of coumarins, and the MaUGT gene may therefore be involved in coumarin biosynthesis. Conclusion It is important to elucidate the autotoxicity and anabolic mechanisms of coumarins to create new germplasms that produce fewer autotoxic substances.

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Section: Survey Methodologymentioning

confidence: 99%

Anabolic metabolism of autotoxic substance coumarins in plants

Wu,

Shi,

Zhang

et al. 2023

PeerJ

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“…Among these workflows, Autodock Vina is a widely used software that uses sophisticated gradient optimization algorithms and has significantly better accuracy of binding mode prediction (Trott & Olson, 2009). Moreover, Autodock Vina has been routinely used in glycoscience field particularly to identify substrate binding regions in the case of glycosyl hydrolases (GHs) and Glycosyltransferases (GTs) (Alsina et al, 2021; Bandi et al, 2021; Pozzo et al, 2014; Xu et al, 2021).…”

Section: Introductionmentioning

confidence: 99%

Machine learning assisted ligand binding energy prediction forin silicogenerated glycosyl hydrolase enzyme combinatorial mutant library

Guranovic

Kumar

Bandi

et al. 2022

Preprint

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Molecular docking is a computational method used to predict the preferred binding orientation of one molecule to another when bound to each other to form an energetically stable complex. This approach has been widely used for early-stage small-molecule drug design as well as identifying suitable protein-based macromolecule residues for mutagenesis. Estimating binding free energy, based on docking interactions of protein to its ligand based on an appropriate scoring function is often critical for protein mutagenesis studies to improve the activity or alter the specificity of targeted enzymes. However, calculating docking free energy for a large number of protein mutants is computationally challenging and time-consuming. Here, we showcase an end-to-end computational workflow for predicting the binding energy of pNP-Xylose substrate docked within the substrate binding site for a large library of combinatorial mutants of an alpha-L-fucosidase (TmAfc, PDB ID- 2ZWY) belonging to Thermotoga maritima glycosyl hydrolase (GH) family 29. Briefly, in silico combinatorial mutagenesis was performed for the top conserved residues in TmAfc as determined by running multiple sequence alignment against all GH29 family enzyme sequences downloaded from an in-house developed Carbohydrate-Active enZyme (CAZy) database retriever program. The binding energy was calculated through Autodock Vina with pNP-Xylose ligand docking with energy minimized TmAfc mutants, and the data was then used to train a neural network model which was also validated for model predictions using data from Autodock Vina. The current workflow can be adopted for any family of CAZymes to rapidly identify the effect of different mutations within the active site on substrate binding free energy to identify suitable targets for mutagenesis. We anticipate that this workflow could also serve as the starting point for performing more sophisticated and computationally intensive binding free energy calculations to identify targets for mutagenesis and hence optimize use of wet lab resources.

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Glycosyltransferases: Mining, engineering and applications in biosynthesis of glycosylated plant natural products

Bai

Tan

et al. 2022

Synthetic and Systems Biotechnology

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Molecular cloning and biochemical characterization of a new coumarin glycosyltransferase CtUGT1 from Cistanche tubulosa

Cited by 3 publications

References 37 publications

Anabolic metabolism of autotoxic substance coumarins in plants

Anabolic metabolism of autotoxic substance coumarins in plants

Machine learning assisted ligand binding energy prediction forin silicogenerated glycosyl hydrolase enzyme combinatorial mutant library

Glycosyltransferases: Mining, engineering and applications in biosynthesis of glycosylated plant natural products

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