Abstract
Lithospermum erythrorhizon produce and highly accumulate a red naphtoquinone pigment shikonin in the root epidermis. A key regulatory enzyme of shikonin biosynthesis is p-hydroxybenzoate(PHB):geranyltransferase (LePGT), which belongs to membrane-bound prenyltransferase family involved in CoQ biosynthesis. LePGT shows strict substrate specificity for geranyl diphosphate (GPP) unlikely to other members of this enzyme family. LePGT composed of 307 amino acids has 9 transmembrane domains, which was functionally expressed in yeast, but the protein yield was very low. In this study we have expressed LePGT in Baculovirus system for the purpose of X-ray crystallographic analysis. The recombinant LePGT expressed in Sf9 have shown much higher specific activity and high protein yield than those of in yeast, which has been solubilized retaining the enzymatic function. We have further established the purification protocol using His-tag attached LePGT.
