The importance of these aromatic NCIs is further supported by the lack-of-function observed when Y177 is mutated to a non-aromatic residue. Y177A/D/L mutants did not afford products with any substrate, with the exception of Y177A with 1a. Interestingly, the differential alkene geometry of 1a (compared to 2a-4a) positioned the substrate alkene substituent on the opposite side of the active site such that the ligand did not necessitate residue 66 repositioning, as revealed by IFD (Fig. ).