Identification and Characterization of Bifunctional Drimenol Synthases of Marine Bacterial Origin.

Natural drimane-type sesquiterpenes, including drimenol, display diverse biological activities. These active compounds are distributed in plants and fungi; however, their accumulation in bacteria remains unknown. Consequently, bacterial drimane-type sesquiterpene synthases remain to be characterized. Here, we report five drimenol synthases (DMSs) of marine bacterial origin, all belonging to the haloacid dehalogenase (HAD)-like hydrolase superfamily with the conserved DDxxE motif typical of class I terpene synthases and the DxDTT motif found in class II diterpene synthases. They catalyze two continuous reactions: the cyclization of farnesyl pyrophosphate (FPP) into drimenyl pyrophosphate and dephosphorylation of drimenyl pyrophosphate into drimenol. Protein structure modeling of the characterized DMS (AsDMS) suggests that the FPP substrate is located within the interdomain created by the DDxxE motif of N-domain and DxDTT motif of C-domain. Biochemical analysis revealed two aspartate residues of the xxE motif that might contribute to the capture of the pyrophosphate moiety of FPP inside the catalytic site of AsDMS, which is essential for efficient cyclization and subsequent dephosphorylation reactions. The middle aspartate residue of the DxTT motif is also critical for cyclization. Thus, AsDMS utilizes both motifs in the reactions. Remarkably, the unique protein architecture of AsDMS, which is characterized by the fusion of a HAD-like domain (N-domain) and a terpene synthase β domain (C-domain), significantly differentiates this new enzyme. Our findings of the first examples of bacterial DMSs suggest the biosynthesis of drimane sesquiterpenes in bacteria and shed light on the divergence of the structures and functions of terpene synthases.