Molecular breeding of carotenoid biosynthetic pathways

The burgeoning demand for complex, biologically active molecules for medici ne, materials science, consumer products, and agrochemicals is driving effo rts to engineer new biosynthetic pathways into microorganisms and plants. W e have applied principles of breeding, including mixing genes and modifying catalytic functions by in vitro evolution, to create new metabolic pathway s for biosynthesis of natural products in Escherichia coli. We expressed sh uffled phytoene desaturases in the context of a carotenoid biosynthetic pat hway assembled from different bacterial species and screened the resulting library for novel carotenoids. One desaturase chimera efficiently introduce d six rather than four double bonds into phytoene, to favor production of t he fully conjugated carotenoid, 3,4,3',4'-tetradehydrolycopene. This new pa thway was extended with a second library of shuffled lycopene cyclases to p roduce a variety of colored products. One of the new pathways generates the cyclic carotenoid torulene, for the first time, in E. coli. This combined approach of rational pathway assembly and molecular breeding may allow the discovery and production, in simple laboratory organisms, of new compounds that are essentially inaccessible from natural sources or by synthetic chem istry.