?-amino acids are an important motif found in many small molecule and large ?-peptide drugs. We endeavor to develop a general method for synthesizing these cores. The focus of this proposal is development of a new method for ?-amino acid synthesis using metalloenzymes to catalyze a C-H amination. Currently there are no known C-H amination catalysts found in nature. We propose evolving the existing machinery of p450 CYP102A1 (p450BM3), an effective catalyst for C-H hydroxylation, to perform a C-H amination at the ?-position of carboxyl groups. Previous studies have demonstrated this enzyme will introduce oxygen into a variety of substrates with O2 as the stoichiometric oxidant. We propose using azides as a nitrogen based oxidant for amination.
Our specific aims are: (1) To evolve existing p450BM3 to aminate esters with long alkoxy- acid linkages to mimic the native substrate for p450BM3;then extend this method to simple esters using directed evolution;(2) Aminate alkyl acids using directed evolution in concert with a hydrogen bonding additive.
These aims will be initiated using the large number of existing p450BM3 mutants in the Arnold Lab followed by further rounds of evolution. We will used a click reaction with a fluorescent alkyne to determine the degree of azide consumption in order to accelerate the screening process. All starting materials can be rapidly prepared or purchased from commercial vendors.
?-amino acids are an important motif found in numerous drug candidates. Further application of this class of molecule is dependent on a robust synthesis. The proposed research will make new variants of this scaffold accessible, which should lead to new and more effective medicinal targets.
|Prier, Christopher K; Hyster, Todd K; Farwell, Christopher C et al. (2016) Asymmetric Enzymatic Synthesis of Allylic Amines: A Sigmatropic Rearrangement Strategy. Angew Chem Int Ed Engl 55:4711-5|
|Farwell, Christopher C; Zhang, Ruijie K; McIntosh, John A et al. (2015) Enantioselective Enzyme-Catalyzed Aziridination Enabled by Active-Site Evolution of a Cytochrome P450. ACS Cent Sci 1:89-93|
|Hyster, Todd K; Farwell, Christopher C; Buller, Andrew R et al. (2014) Enzyme-controlled nitrogen-atom transfer enables regiodivergent C-H amination. J Am Chem Soc 136:15505-8|
|Farwell, Christopher C; McIntosh, John A; Hyster, Todd K et al. (2014) Enantioselective imidation of sulfides via enzyme-catalyzed intermolecular nitrogen-atom transfer. J Am Chem Soc 136:8766-71|