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Expression, purification and structural characterization of resveratrol synthase from Polygonum cuspidatum
Mei Dai1, Daopeng Yuan1, Yangmei Lei2
1Biological Physics Laboratory, College of Science, Beijing Forestry University, Beijing, 100083, China.
Protein Expression and Purification
|November 22, 2021
Summary
Resveratrol synthase (RS) from Polygonum cuspidatum, named PcRS1, was successfully expressed and purified. Structural analysis revealed enhanced catalytic capacity for resveratrol synthesis.
Area of Science:
- Biochemistry
- Molecular Biology
- Plant Science
Background:
- Polygonum cuspidatum is a medicinal plant rich in resveratrol.
- Resveratrol synthase (RS) is key to resveratrol synthesis and highly expressed in P. cuspidatum stems.
Purpose of the Study:
- To elucidate the molecular basis of resveratrol synthesis by PcRS1.
- To characterize the structure and function of the PcRS1 protein.
Main Methods:
- Amplification and sequencing of the RS gene.
- Expression and purification of recombinant PcRS1 protein in E. coli.
- Ni-NTA chromatography, SDS-PAGE, SEC, and DLS for protein characterization.
- Structural prediction and analysis of PcRS1-ligand complexes.
Main Results:
- The PcRS1 gene encodes a 389-amino acid protein with a molecular weight of 42.4 kDa.
- Highly pure and stable soluble PcRS1 protein was obtained.
- Enzyme activity was verified by detecting reaction products at 303 nm.
- Structural analysis revealed an optimal active site configuration, enhancing catalytic capacity.
Conclusions:
- PcRS1 possesses enhanced catalytic efficiency for resveratrol synthesis due to its structural configuration.
- This study provides insights into the structural evolution of resveratrol synthases.
