Molecular cloning and characterization of the CHS gene family in turmeric (Curcuma longa Linn.)

Three chalcone synthase (CHS) genes were isolated from Curcuma longa Linn. using TAIL-PCR. ClCHS1 and ClCHS2 were 1,460 and 1,407 bp in length, respectively, containing 1,191 bp open reading frame (ORF) that encodes 396 amino acids, whereas ClCHS3 was 1,394 bp in length containing 1,170 bp ORF that encodes 389 amino acids. The structure of all three genes comprise two exons and one intron which are consistent with the other CHS gene family. Southern blot analysis using a ClCHS conserved fragment revealed the ClCHS genes belong to a gene family. Phylogenetic analysis showed the three putative ClCHS proteins to be closely related to DCS (diketide-CoA synthase) protein, a product of CHS-like gene in C. longa, which condenses malonyl-CoA with feruloyl-CoA or coumaroyl-CoA as the substrate in curcuminoid synthesis. These results suggest that the interaction of substrate and enzyme between the three putative ClCHS proteins and DCS might be highly similar. Homology modeling and docking analysis were consistent, indicating that the same substrate (coumaroyl-CoA) can be used in the putative ClCHS1 and ClCHS2 proteins. However, the putative ClCHS3 protein seems to have seven amino acids deletion in a loop involved in the binding site formation, suggesting that the binding site with coumaroyl-CoA might be altered.