(5cz) Enzymatic Synthesis of Simvastatin

Natural products are small molecules found in diverse natural sources such as bacteria, fungi, plants and so on. They are produced by dedicated biosynthetic pathways using enzymes that possess exquisite catalytic power. Enzymes discovered from these pathways collectively constitute a versatile molecular toolbox for use in drug discovery and drug synthesis. Simvastatin (Zocor), one of the best selling drug in the united states with annual sales ~$5 billion before the expiration of the patent on 2007, is a cholesterol-lowering drug due to its potent inhibitory activity towards the rate-limiting step of cholesterol biosynthesis. Currently simvastatin was semisynthesized from lovastatin, a natural product derived from Aspergillus terreus.

We enzymatically synthesized simvastatin by using of an acyl-transferase, LovD (46 kDa), which is involved in the lovastatin biosynthetic pathway. We first demonstrated that LovD has broad substrate specificity. We then developed an Escherichia coli based whole-cell platform, which can readily synthesize simvastatin from easily accessible substrates. The new process avoided the currently used multiple-step organic synthesis of simvastatin and the final yield was much higher. Later, multiple techniques, including screening of the substrates, metabolic engineering of E. coli, protein engineering of LovD, etc., were carried out to optimize the process All of these significantly improved the whole cell biocatalytic process. High cell density fermentation opens the door for industrial production of simvastatin at lower cost. Besides that, to understand the fundamental details of naturally happened lovastatin biosynthesis, the megasynthase LovF from the same pathway was expressed and purified from Saccharomyces cerevisiae. We showed that protein-protein interactions between LovF and LovD facilitated highly efficient biosynthesis of lovastatin. To understand the basis of LovD catalyzed reaction, X-ray crystallography was employed, which provided atomic resolution of the mechanism of catalysis, substrate and product binding, and a likely basis of beneficial mutations.