7. Project Summary The work described in this proposal aims to advance biocatalytic tools for use in the synthesis of medicinally valuable molecules. We will develop new reaction mechanisms for pyridoxal-dependent enzymes to enable the facile synthesis of non-canonical amino acids through photoexcitation of pyridoxal intermediates involved in the catalysis of threonine aldolase and serine hydroxymethyltransferase. The first goal is to photoexcite the PLP external aldimine enabling it to function as a photoacid. This mechanism will allow these enzymes to use a broader set of 𝛼-, 𝛽-, and 𝛾-amino acids as pronucleophiles. The second goal is to excite the PLP-quinonoid to access a more reducing excited state. This species can reduce alkyl halides, redox-active esters, and cyanoheterocyles to form radicals that can recombine with the PLP-quinonoid radical to produce various non-canonical amino acids. Collectively, these studies will provide a new tool for the synthesis of non- canonical amino acids and serve as the foundation for a new area in photoenzymatic catalysis. The methods and the goals proposed in the Specific Aims can streamline the synthesis of biological probes and drug targets, creating a significant benefit to human health and associated biomedical sciences.

8. Project Narrative This proposal will develop novel mechanisms for PLP-dependent enzymes enabling the streamlined synthesis of non-canonical amino acids. This work provides a solution to the long-standing challenge of preparing molecules with polar functional groups without protecting and activating groups. These new reactions and catalysts will provide biomedical sciences with new tools to accelerate the discovery of new medicines, creating a significant benefit to human health.