Characterization of the aminocoumarin ligase SimL from the simocyclinone pathway and tandem incubation with NovM,P,N from the novobiocin pathway

Simocyclinone D(8) consists of an anguicycline C-glycoside tethered by a tetraene diester linker to an aminocoumarin. Unlike the Antibiotics novobiocin, clorobiocin, and coumermycin A(1), the phenolic hydroxyl group of the aminocoumarin in simocyclinone is not glycosylated with a decorated noviosyl moiety that is the pharmacophore for targeting Bacterial DNA gyrase. We have expressed the Streptomyces antibioticus simocyclinone Ligase SimL, purified it from Escherichia coli, and established its ATP-dependent amide bond forming activity with a variety of polyenoic acids including retinoic acid and fumagillin. We have then used the last three Enzymes from the novobiocin pathway, NovM, NovP, and NovN, to convert a SimL product to a novel novobiocin analogue, in which the 3-prenyl-4-hydroxybenzoate of novobiocin is replaced with a tetraenoate moiety, to evaluate Antibacterial activity.