This project is directed at the development of new methods for use of enzymes as catalysts in the synthesis of organic and biological compounds. One area of work will be the development of new methods in the enzymatic synthesis of analogs of coenzyme A (CoA). This methodology will be used in the synthesis of compounds useful in probing the mechanism and structure of enzymes which utilize CoA or a CoA ester as substrate. The CoA and CoA ester utilizing enzymes include many enzymes of pharmaceutical and medical importance. Studies facilitated by CoA analogs prepared in this project will be valuable in providing new insights into mechanisms and means of inhibition of these enzymes. A truncated CoA analog will be prepared and used for the activation of simple thioesters as substrates for CoA ester utilizing enzymes by a reversible linkage strategy. This technology will be employed in the synthesis of compounds useful in the preparation of a variety of pharmaceutical agents. In related work, a method for the synthesis of analogs of acyl carrier protein (ACP) will be developed. This synthesis will utilize the enzyme holo-ACP synthase and CoA analogs previously prepared in this lab. ACP has a function analogous to that of CoA, and is utilized in the biosynthesis of fatty acids and polyketide antibiotics. ACP analogs will serve as tools for deciphering mechanistic issues in these enzymes. This information may be useful in design of inhibitors of fatty acid biosynthesis, a potential target for anti-cancer therapy. This information may also be valuable in learning how to redirect polyketide biosynthesis to make analogs of the natural polyketide antibiotics, which may have useful biological activity. Work will also be conducted in the enzymatic resolution of carboxylic acids having a chiral center at the alpha-carbon. This work will make use of thioesters as substrates for lipases and other hydrolases under conditions in which the substrate is induced to continuously racemize, by deprotonation and reprotonation at the alpha-carbon. Coupled with selective hydrolysis of the desired enantiomer, this will permit quantitative conversion of racemic starting material into one isomer of product. Products prepared will include compounds of known pharmaceutical value and potential synthons for the preparation of other biologically active compounds.
