To further probe the scope of accepted cyclic substrates, we initially challenged the three G-type HHDHs with the model substrates 3-10 using azide as the nucleophile (Figure ). In addition to the known substrates 1 and 2, all enzymes displayed activity with cyclopentene oxide (3), the -methylated epoxide 4 and a cyclic epoxide bearing a distal vinyl group (5) with most reactions reaching nearly full conversion after 24 h. In contrast, electronically challenging substrates such as the ketone 6 or tetrahydrofurane 7 were not converted. Similarly, incubation of the highly sterically demanding bridged tricycle 8 and the octa-and decacycles 9 and 10 with the three enzymes gave no conversion to the corresponding azidoalcohols. Following these initial tests with purified enzyme, we additionally conducted preparative-scale reactions with 3-5, azide and whole cells harboring HheG, which yielded azidoalcohols 3a-5a in moderate to good yield. Analysis by chiral GC and NMR revealed that all products were obtained as mixtures of regioisomers with low to moderate enantiomeric excess but generally high total turnover numbers (TTNs, Figure ). For instance, the azidolysis of 4 with 0.02 mol% HheG3 proceeds with full consumption of the starting material and a significant preference for attack at the sterically less hindered site to give azidoalcohol 4a. However, background reactivity of the nucleophile in buffered solution consistently affords a mixture of regioisomers (Figure ). TTNs were determined by analysis of crude reaction mixture by achiral GC. Please see the Supplementary Information for the full experimental details and analytical procedures.