Doug Matje
Research Abstract
Billions of years of selective pressures have evolved enzymes to be the most powerful, precise catalysts on the planet. The repertoire of products produced gives us all of the natural materials we use, half of the medicines we consume and all of the food we eat, along with regulating and directing nearly every cellular process. For nearly every organic natural product created, numerous inorganic, and many synthetic products only recently produced exclusively in a laboratory, there exists an enzyme able to degrade the molecule. However, nearly all the catalytic capacity of enzymes has been evolved in-vivoas a response to environmental pressures. Despite no lack of effort on the part of bioengineers, the generation of novel specificity and function in enzymes has been limited. The slow progress on the potentially unlimited chemical power of enzymes is due to many factors, including a lack of full understanding of the incredibly complex and dynamic nature of even a single macromolecule, a limited repertoire of chemical functionality in the natural amino acids, and lack of high-throughput screens needed to assess the vast amino acid sequence space available to even a relatively small enzyme. My interests lie in using existing techniques, such as the use of unnatural amino acids and directed evolution, as well as developing new methodologies, to develop new enzyme activities to take advantage of the incredible power of nature's catalysts.
My current project involves the insertion of an unnatural amino acid into one of our lab's most well characterized enzymes, MHhaI, a bacterial cytosine DNA methyltransferase. Based upon the detailed structural knowledge of the active site and amino acid residues important in determining specificity, catalysis, and for triggering a large conformational change that leads to catalysis, my project involves inserting a citrulline in place of a arginine residue to change the specificity of the enzymes cognate site from GCGC to ACGC (methylated cytosine shown in bold). We then hope to use directed evolution to improve the potentially debilitating consequences for catalysis generated by this insertion using in-vitro compartmentalization. It is our hope that this research will lead to the ability to fine tune specificity of DNA restriction/modification systems for use in biomedical and nanobiotechnology applications such as DNA SMILing.
Papers
Research Support
Presentations
Contact
Reich Lab Contact Info