Thousands of terpenes and terpenoid derivatives are found throughout nature and are involved in diverse biosynthetic and metabolic pathways such as cholesterol biosynthesis in humans and paclitaxel (Taxol) synthesis in the Pacific yew. Notably, many terpenoids have been used as medicinal agents since the times of antiquity due to their analgesic, antibiotic, and antifungal properties. In spite of the universal importance of this family of natural products for human health, it is remarkable that the three-dimensional structures of terpenoid cyclases have only recently been reported. Terpenoid cyclases (aka synthases) catalyze the specific cyclization of a common allylic pyrophosphate substrate, such as farnesyl diphosphate, into one of 300+ possible products. Enzymological studies of terpenoid cyclases indicate that the enzyme plays a critical role in channeling the precise substrate (plus/or intermediate) conformation to lead to the formation of one exclusive product. Thus, the terpenoid synthases comprise an exciting class of biosynthetic enzymes from both the biological and the chemical perspectives, and here we address the dearth of structural information on this enzyme class.
We aim to dissect structure-function relationships in the active site of the sesquiterpene cyclase, pentalenene synthase. This terpenoid synthase was originally isolated from Streptomyces UC5319 and is important in the biosynthesis of the pentalenolactone family of antibiotics. Additionally, we aim to determine the crystal structures of two other sesquiterpene cyclases: aristolochene synthase, which catalyzes the stereospecific formation of a precursor to a family of antifungal compounds; and trichodiene synthase, which catalyzes the formation of the precursor to the antibiotic trichcethin. Unexpected similarities between the structure of pentalenene synthase and the prenyl transferase, farnesyl diphosphate synthase, are suggestive that the terpenoid cyclases possess a universal active site structure specifically adapted in each case as a unique template to channel the substrate binding conformation. We will further define structure- function relationships in these terpenoid cyclases by determining the crystal structures of rationally designe variants and enzyme-inhibitor complexes. This work is prerequisite for the structure-based redesign of these enzymes to generate natural and unnatural products of potential medicinal importance.
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