Since the variant T123M was either not present in our previous library screen for the T123 position in HheG or did not exhibit an increased thermostability, the origins of HheG3's increased stability currently remain elusive. In accordance with their 𝑇 𝑚 values, HheG and HheG3 displayed much higher stability under typical reaction conditions, as both enzymes retained measurable activity in the azidolysis of 1 after 24 h of incubation, while HheG2 was rapidly inactivated (Figure ).