Previous work using random mutagenesis and screening already yielded mutants F26L and R69H that, compared to F26H and R69K described herein, are slightly better catalysts for transglycosylation (1.2-fold). However, the current strategy is less labour-intensive, as it circumvents large library screening. In this respect, it is remarkable that the variants H240N and H240F display different yields and regioselectivities (Table ), thus illustrating the value of further probing and fine-tuning of hotspots identified using our strategy.