Enzymes isolated from microbes play significant roles in daily life, ranging from photosynthesis and oxygen production by algae to yogurt formation by Lactobacilli and Lactococci bacteria. The proposed research will investigate a unique, poorly-understood mechanism for the methylation of unreactive carbon and phosphorus atoms used by enzymes in Streptomyces, a large genus of beneficial bacteria that naturally produce many antibiotic, anticancer, and herbicidal chemicals used commercially. To date, all enzymatic methyl transfer reactions have been characterized as nucleophilic substitution reactions. However, the substrates for these Streptomyces methyltransferases are not amenable to nucleophilic activation, suggesting the need for and use of another catalytic mechanism. Biochemical, spectroscopic, and synthetic organic techniques will be used to determine whether these enzymes catalyze methyl transfer using a four-iron, four-sulfur cluster to generate organic radicals from S-adenosyl-L-methionine (SAM) and methylcobalamin, a derivative of vitamin B12. Understanding the mechanism of these reactions will provide a framework for engineering these and related enzymes to produce novel carbon-carbon and/or carbon-phosphorus-carbon bonds. The enzymes, cofactors, and microbes studied in this research will be linked to an educational outreach program targeting K-5th grade children of the rural, agricultural Palouse region, which is 300 miles from the nearest metropolitan area. In partnership with the Palouse Discovery Science Center, interactive presentations focusing on beneficial microbes and enzyme chemistry will be presented to families and classes throughout the area. Giveaways will be combined with assessments to improve retention and obtain feedback for improvements. Similar presentations incorporating experimental results will also be made available in an accessible online format to reach a larger audience. The ultimate goals of this proposal are to improve overall understanding of microbial enzyme chemistry and encourage children to pursue future careers in microbiology and/or biochemistry.
