For determination of specific activities in epoxide ring opening with epoxide substrates 1, 2, 3, 4, 5, and 6 using azide as nucleophile, reactions of 1 mL total volume contained 10 mM epoxide, 20 mM azide, and 10-400 µg mL -1 purified enzyme in 2 mM MOPS•SO4 pH 7.0. Reactions were incubated at 22 °C (HheG wild type and mutants) or 30 °C (HheC wild type and mutants) with shaking at 900 rpm in a ThermoMixer C from Eppendorf (Hamburg, Germany). Samples of each 100 µL were taken after 30-360 s and transferred to a 96-well plate containing already 100 µL quenching solution (40 µg mL -1 BTB in 100% methanol) per well. Afterwards, absorbance at 616 nm and 499 nm was measured using a ClarioStar microplate reader. Activities were calculated using equations S8, 10, 15, 17, 18 in the supplementary. This assay was also used for the determination of kinetic parameters in epoxide ring opening reactions. For kinetic measurement of HheC and its mutants, reactions were carried out in 2 mM MOPS•SO4 using 1-150 mM epichlorohydrin (1) while keeping the azide concentration fixed (60 mM for HheC WT, T7S, F12Y, 100 mM for HheC G14A) or using 1-300 mM azide while keeping the concentration of epoxide 1 constant at 100 mM. Reactions were incubated at 30 °C. Samples were taken after 30-360 s to ensure determination of initial velocities. For HheG and its mutants, reactions were performed in 2 mM MOPS•SO4 using 1-150 mM cyclohexene oxide (2) while keeping the azide concentration fixed (60 mM for HheG WT, T13S, Y18F, 100 mM for HheG G20A) or using 1-300 mM azide while keeping the concentration of epoxide 2 constant at 100 mM. Reactions were incubated at 22 °C. Samples were taken after 30-360 s to ensure determination of initial velocities. Analysis of the resulting kinetic data was performed as previously described for specific activity calculation and kinetic data was fitted using the Hillequation ( ) in Origin Pro2021 (see Figures ).