Tailoring enzymes isolated from the biosynthetic pathways of structurally unique compounds are of special interest because they offer a novel mechanism for synthetically modifying therapeutic lead compounds with diverse functional groups. It is the goal of this study to isolate and characterize proteins required for a novel tailoring reaction in the marine cyanobacteria, Lyngbya majuscula. By using the combined techniques of biochemical fractionation and molecular biology, we propose to isolate and characterize the enzyme(s) required for the trichlorination of barbamide. Our goal will be to use the recombinant enzymes as a tool for optimizing the activity of therapeutic lead compounds. After isolation and microsequencing of candidate enzymes, halogenating enzyme(s) will be cloned and efforts made to apply the activity of the enzymes to the tailoring of other natural products with therapeutic relevance. Derivatization reactions will be generated by transforming biosynthetically active species of bacteria and cyanobacteria with the unique barbamide chlorinating enzymes. Alternatively, the incorporation of trichloromethyl groups into therapeutic lead compounds will be accomplished in vitro using enzyme-mediated reactions or via feeding experiments with chlorinated substrates that are generated using purified recombinant enzymes.
