Recombination and Screening of Putative Grapefruit Glucosyltransferase 4 in Pichia pastoris

Flavonoids are a group of plant secondary metabolites that are vital to the cell systems of plants. The intake of these chemicals is advantageous to animals for their antioxidant properties that affect the function of immune and inflammatory cells. The bitter taste of grapefruit (Citrus paradisi) and other citrus species is caused by the accumulation of glycosylated flavonoids. Glucosyltransferases (GTs) are enzymes that add glucose moieties to a carbon or hydroxyl group of natural products. The function of a putative secondary product GT clone was tested. In previous research, putative GT 4 was cloned into a pCD1 modified pET expression system, heterologously expressed in E.coli, and screened for activity with a few substrates; little GT activity was found. Issues of protein localized to inclusion bodies in bacteria were addressed. PGT 4 is being heterologously expressed in yeast (Pichia pastoris) to allow for protein production and analysis. PGT 4 was screened for GT activity with different flavonoid subclass representatives and simple phenolics