Discovery and application of halogenases for medicinal chemistry

Project Summary Halogen atoms (F–, Cl–, Br–, and I–) are incredibly versatile components of both naturally produced and synthetic compounds. Over half of the pharmaceuticals on the market in the United States contain at least one halogen atom, and likely many more compounds involve halogen atoms in their synthesis. Chemical methods for installing halogen atoms typically generate one or more undesired side products and require reagents that have adverse effects for human and environmental health. Methods improving both halogenation selectivity and sustainability are therefore highly desired. My lab approaches these general needs by studying enzymes to complement synthetic chemical methods. Specifically, we focus on halogenases capable of installing halogen atoms onto organic compounds with precise selectivity in an environmentally-benign manner. In the area of medicinal chemistry, halogenases have the potential to transform the landscape of environmentally-conscious catalysis, but shortcomings related to enzyme stability, scope of substrate preference, and reaction kinetics have limited their broad application thus far. To combat these known limitations, my group combines bioinformatics, natural product biosynthetic logic, and enzymology to identify novel enzymes in publicly available genomic data with biocatalytic potential. Using these techniques, we have identified a new family of halogenases that is phylogenetically unrelated to known halogenases and possesses superior kinetic properties, enhanced stability, and broad substrate scope. Within this family exist two distinct chemoselectivity preferences where one subsection of the family targets indole-containing molecules such as the amino acid L-tryptophan and the other selectively installs halogen atoms onto terminal alkynes to generate haloalkynes, a reaction that was previously unknown in enzyme chemistry. The initial discovery of these powerful biocatalysts inspires the focus of this proposal, which places emphasis on expanding our fundamental understanding of the biochemistry of this new halogenase family and developing both scalable and high throughput biocatalytic platforms to leverage the potential of these enzymes in medicinal chemistry. In the period of the next five years, we aim to break new ground in halogenation chemistry and biocatalysis, develop robust and generalizable methodologies to apply to other enzyme systems in our lab, and identify the next phase of biocatalytic innovation in our program through natural product discovery. Through the highly interdisciplinary research described in this proposal, members of the Lukowski lab are trained in enzymology, natural products chemistry, biosynthesis, analytical chemistry, biocatalysis, organic chemistry, microbiology, synthetic biology, marine science, and bioinformatics, enabling challenging problems to be tackled and preparing the next generation of researchers for careers in industry, academia, government, and the private sector.