Isolation and Characterization of Chalcone Isomerase (CHI) Gene from Boesenbergia rotunda

Abstract

Chalcone isomerase (CHI; E.C. 5.5.1.6) is an enzyme that plays critical roles in the flavonoid biosynthesis pathway and plant secondary metabolism. Although CHI genes have been widely studied in various higher plants, to date there are no studies about CHI genes in Boesenbergia rotunda. The CHI gene (747 bp) was isolated from different parts of the plant, including rhizomes, leaves and flowers of B. rotunda. The isolated B. rotunda CHI, designated BrCHI, contained one intron and two exons and encoded a peptide of 248 amino acid with a predicted molecular mass of 26.2 kDa and pI of 5.03. The deduced amino acid sequence of BrCHI shares 63–65% identity with CHI from other plants within the order of Zingiberales such as banana and canna lily. Sequence alignment showed that the coding region of the BrCHI gene is highly similar to CHI genes from other plant species and has four conserved amino acid residues (Thr53, Tyr112, Asn119, and Ser191) which are essential for CHI function. Phylogenetic analysis has clustered BrCHI together with other CHIs belonging to CHI type I. Recombinant BrCHI protein was successfully expressed in E. coli using the pMAL-c4X vector cloning system. In vitro enzyme activity, assayed by HPLC, indicated that recombinant BrCHI could catalyse the formation of naringenin from naringenin chalcone. This study demonstrates the ability of bacterial host to overexpress BrCHI gene isolated from B. rotunda to produce functional BrCHI that successfully converts naringenin chalcone to naringenin