Mechanistic Studies of C-X Bond Formation Chemistries

With the support of the Chemistry of Life Processes (CLP) program in the Division of Chemistry, Professor Pinghua Liu of Boston University is studying the structure-function relationship of C-S bond formation using enzymes in ovothiol biosynthesis as the model system. Sulfur represents ~1% of the cell dry weight and sulfur-containing molecules play important roles in biological systems. Biosyntheses of sulfur-containing natural products, especially by selective C-H activation, are at the frontiers of mechanistic enzymology and biocatalysis. The proposed structure-function relationship studies of ovothiol biosynthetic enzymes are important addition to this field. The proposed work integrates tools from organic/bio-synthesis, structural biology, kinetics, spectroscopies, and enzyme engineering, which is an excellent platform to train STEM students. Reactions studied in this work may also be applied to improve ovothiol industrial production processes due to its biological activities. Ovothiol biosynthetic enzymes (OvoA) were selected in this study due to several of their unique features: 1) crystal structures have been obtained; 2) some key intermediates have been trapped; and 3) they are flexible in substrate selectivity. The C-S bond formation reactions performed by OvoA are distinct from current literature examples. With this award, the proposed structure-function relationship studies seek to provide mechanistic understanding on factors modulating the partitions among several different reaction pathways in these enzymes. These studies include characterizing factors governing the C-S bond formation regioselectivity. Moreover, pre-steady state and spectroscopic characterization of the OvoA enzyme, especially factors that might affect the kinetics and properties of the S = 1 Fe(IV) species will be carried out. Finally, biochemical, kinetic, and spectroscopic characterization of OvoA’s cysteine dioxygenase activity will be performed. The central educational objective of this proposal is to train our students using an interdisciplinary approach and then apply knowledge gained in basic research to solve real world problems. This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.