Pterostilbene production by microorganisms expressing resveratrol O-methyltransferase

Pterostilbene (3,5-dimethoxy-4′-hydroxyl-trans-stilbene)—a derivative of resveratrol—is a natural dietary compound and the primary antioxidant component in berries. Pterostilbene has significant advantages over resveratrol in bioavailability, half-life in the body, cellular uptake, oral absorption and metabolic stability. Here, we expressed the resveratrol O-methyltransferase (ROMT) gene (VvROMT) from grape (Vitis vinifera) in Escherichia coli and Saccharomyces cerevisiae and confirmed its specific ability to catalyze the production of pterostilbene from resveratrol. By co-expressing an additional two genes from the resveratrol biosynthetic pathway—4-coumarate CoA-ligase (4CL) and stilbene synthase (STS)—a large amount of pterostilbene was produced, with a trace amount of pinostilbene detected. To understand the molecular basis of the catalytic activity, four key amino acid residues were identified in a 3D-model of VvROMT and mutagenized and assayed for augmented catalytic activity. Our results demonstrate the potential utility of the engineered microorganisms for pterostilbene production and provide protein engineering targets that will hopefully lead to increased activity of the ROMT enzyme.