The Discovery of Germacradienol Synthase: Construction of Genetically-Engineered Strain, Glycosylated Modification, Bioactive Evaluation of Germacradienol.

Germacradienol is a main precursor in the biosynthesis of geosmin-type terpenes by a variety of microbes, but its biological activities are still unknown. In the biosynthetic mechanism study of an antifungal degraded sesquiterpenoid (1 beta,4 beta,4a beta,8a alpha)-4,8a-dimethyloctahydronaphthalene-1,4a(2H)-diol (5) with a geosmin scaffold, the germacradienol synthase B7C62_00490 was identified. To exploit the synthetic potential of the enzyme to create germacradienol, engineered strains were constructed by introducing key synthases of farnesyl diphosphate, germacradienol synthase B7C62_00490 and glycosyltransferase UGT73C5 in Escherichia coli BL21(DE3). Germacradienol (1) and the novel glycosylated derivate germacradienyl-11-O-beta-D-glucopyranoside (3) were successfully obtained from engineered strains. The cytotoxic activity against nine human cancer cell lines and antimicrobial activities against a panel of bacteria and fungi of germacradienol analogs derived from engineered strains were evaluated. Germacradienol demonstrated multiple biological activities, including broad antimicrobial activities with MIC values ranging from 12.5 to 25.0 mu g/mL and cytotoxic activities with IC50 values ranging from 21.0 to 83.5 mu M. However, the glycosylated germacradienol was inactive.