Isolation, cloning and bioinformatics analysis of β-amyrin 11-oxidase coding sequence from licorice

Abstract

Licorice is the roots and stolons of Glycyrrhiza uralensis which have several chemical compounds. Triterpene saponins such as glycyrrhizin and glycyrrhetinic acid and flavonoids like liquiritin, isoliquiritigenin and glabron are main compounds detected in liquorice root. The plant's major constituent is a glycyrrhizin. The β-amyrin 11-oxidase catalyzes the sequential two-step oxidation of β-amyrin in C-11 to produce 11-oxo-β-amyrin, a possible biosynthetic intermediate between β-amyrin and glycyrrhizin. In this study, the total RNA was extracted from licorice roots and cDNA synthesis, then PCR products were cloned into pTZ57R/T vector. Sequencing confirmed piece length of 1482 bp that encodes a protein of 493 amino acid residues. The results of alignment showed 99% similarity to β-amyrin sequence of Glycyrrhiza uralensis. Subcellular studies using Softberry and Psort software showed that the activity of this protein is in endoplasmic reticulum. Moreover the protein has a signal peptide and is targeted to the secretory pathway. The results of phylogenetic tree determined most similar amino acid sequence to the CYP88D subfamily of cytochrome P450. These findings can be used for nucleotide or protein manipulation and transformation.