Stevia plants synthesize steviol glycosides as a bio-chemical defense against some species of insects. Steviol glycosides are synthesized from products of photosynthesis and glucose breakdown in a complex biosynthetic pathway. Steviol – the backbone of steviol glycoside molecules belongs to diterpene group of compounds – which plays a multitude of biological functions in plants.
Production of all diterpene group of compounds shares a common initial biosynthetic pathway. The diterpene compounds have a basic backbone of “poly-isoprene”. Isoprene is a simple organic compound with the following chemical structure -
The structure of poly-isoprene is as follows
One of the simple diterpene with poly-isoprenoid structure is Geranylgeranyl diphosphate – which is a precursor of several compounds in plants including steviol glycoside.
Steviol glycosides are synthesized in two stages. In the first or early stage, Geranylgeranyl diphosphate or GGDP is synthesized from two of its precursors, isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP). Geranylgeranyl diphosphate is the starting point for many biologically important diterpene compounds in plants. In the late stage, Geranylgeranyl diphosphate is converted into steviol glycosides in several steps. The production of isopentenyl diphosphate (IPP) and dimethylallyl diphosphate (DMAPP) is achieved through a very important pathway in plants, which is called 2-C-methyl-D-erythritol-4 phosphate pathway or MEP pathway. Now let us have a closer look at the MEP pathway, and formation of GGDP. All the steps in this pathway are catalyzed by specific enzymes. In absence of those specialized enzymes, the biochemical reactions do not go forward. In the process flow diagram, the enzymes required are shown in green box
Combining of Pyruvate and Glyceraldehyde 3 phosphate into Deoxyxyulose 5 phosphate and its further phosphorylation by Cytidine triphosphate. This is the start of the MEP Pathway
Further phosphorylation by Adenosine Triphosphate (ATP) followed by formation of 2-C-Methyl-D-Erythritol -2,4-Cyclodiphosphate
Formation of Isopentenyl Diphosphate and Dimethylallyl Diphosphate. These are precursors of a variety of isoprenoid compounds in plants. This is the concluding step of MEP pathway
Formation of Geranylgeranyl diphosphate and its cyclization to form (-) Copalyl diphosphate . This is the start of the late phase of Steviol glycoside synthesis
Then Kaurene is produced from Copalayl Diphosphate by an ionization dependent cyclization. Kaurene is then oxidized to kaurenoic acid. The next step is hydroxylation of kaurenoic acid to form Steviol and this is the first committed step for steviol glycoside production. From here, the steviol glycoside biosynthetic pathway takes a diversion from gibberellin (an important plant growth hormone) biosynthesis pathway.
In the next steps, Steviol is progressively glycosilated with the help of Glucosyl Transferase enzymes to form different steviol glycosides.