Metabolization of the artificial secondary metabolite 4-hydroxybenzoate in ubiC-transformed tobacco

Abstract

Transgenic Nicotiana tabacum cell cultures formed a new, artificial secondary metabolite, i.e. 4-hydroxybenzoate (4HB), by expression of the bacterial gene ubiC, which encodes chorismate pyruvate-lyase. 4HB was converted in the cells to two different glucosides, i.e. 4-0-(1-β-D-glucosyl)benzoic acid (4HBOG) and 4HB 1-β-D-glucosyl ester (4HBCOOG). The same metabolization was also observed with exogenously fed 4HB. This glucosylation is catalyzed by constitutively expressed glucosyltransferases which could be detected in cell-free extracts of transgenic and untransformed cells in the same activity. The enzymes forming the two respective glucosides differ in their pH optima, their affinity for 4HB and their regulation. Both in vivo and in cell-free extracts, 4HBOG is the main product at low 4HB concentration, and 4HBCOOG at high 4HB concentration. Whereas 4HBOG is accumulated as an apparently stable secondary metabolite, 4HBCOOG presents a metabolically active form of 4HB. After feeding of [U14C]4HB, a transient accumulation of 4HBCOOG was followed by incorporation of radioactive 4HB into the cell wall, suggesting that 4HBCOOG, but not 4HBOG serves as precursor of cell wall bound 4HB.