Isolation of 4-hydroxy 3-methyl 2-butenyl 4-diphosphate reductase () gene of methyl erythritol diphosphate (MEP) pathway, analysis and differential tissue specific expression in (Burm. f) Nees.

UNLABELLED

The full length 4-hydroxy 3-methyl 2-butenyl 4-diphosphate reductase () gene of MEP pathway was isolated for the first time The ORF with 1404 bp flanked by 100 bp 5'UTR and 235 bp 3'UTR encoding 467 amino acids (NCBI accession number: MK503970) and cloned in pET 102, transformed and expressed in BL21. The protein physico-chemical properties, secondary and tertiary structure were analyzed. The Ramachandran plot showed 93.8% amino acids in the allowed regions, suggesting high reliability. The cluster of 16 ligands for binding site in involved six amino acid residues having 5-8 ligands. The Fe-S cluster binding site was formed with three conserved residues of cysteine at positions C123, C214, C251 of . The substrate HMBPP and inhibitors clomazone, paraquat, benzyl viologen's interactions with were also assessed using docking. The affinity of Fe-S cluster binding to the cleft was found similar to HMBPP. The HPLC analysis of different type of tissue (plant parts) revealed highest andrographolide content in young leaves followed by mature leaves, stems and roots. The differential expression profile of suggested a significant variation in the expression pattern among different tissues such as mature leaves, young leaves, stem and roots. A 16-fold higher expression of was observed in the mature leaves of as compared to roots. The young leaves and stem showed 5.5 fold and fourfold higher expression than roots of . Our result generated new genomic information on which may open up prospects of manipulation for enhanced diterpene lactone andrographolide production in .

SUPPLEMENTARY INFORMATION

The online version contains supplementary material available at 10.1007/s12298-021-00952-0.