Molecular cloning and enzymatic characteristics of chalcone isomerase from Rheum palmatum

Authors: XIAOWEI HUO, GANG ZHANG, MENGMENG LIU

Abstract: Rheum palmatum has been widely developed in traditional Chinese medicine. One of the major active compounds in R. palmatum is flavonoids. Until now, few genes involved in flavonoid biosynthesis have been isolated from this medicinal plant. In this study, a new chalcone isomerase gene (RpCHI) was cloned from an RNA-seq library of R. palmatum. The 714bp RpCHI cDNA was cloned and sequenced. The RpCHI recombinant protein was successfully overexpressed in a prokaryotic expression system and purified to homogeneity by a nickel-nitrilotriacetic acid (Ni-NTA) prepacked column, and the enzymatic activity was investigated. The results have clarified that RpCHI was assembled into the type I CHI group based on phylogenetic analysis. Activity examination revealed that RpCHI exhibits high substrate affinity for naringenin chalcone, and it can also use isoliquiritigenin in relatively low activity. Optimum pH and temperature for enzymatic reaction was determined to be 7.0 and 40 °C, respectively. RpCHI was successfully cloned in the experiment, and the enzymatic activity revealed RpCHI uses various substrates to produce antioxidant flavonoids.

Keywords: Rheum palmatum, chalcone isomerase, sequence analysis, molecular cloning, enzymatic activity

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