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Finally, we challenged the enzymes with
nitrene precursor 1z, which has three sets of
reactive C(sp3)−H bonds. Variants LSb, LSg, and
LSsp3 selectively formed b-lactam 2z, g-lactam
6z, and d-lactam 3z, respectively. The site selec-
tivity shown in this example further showcases
the powerful regiocontrol of the enzymes that is
tunable by directed evolution (Fig. 3D).
We have engineered a “lactam synthase” that
can be tuned by directed evolution to convert
individual substrates into different lactams through
a catalyst-controlled C–H amidation process. Re-
activity trends due to bond strength, inductive
effects, steric accessibility, or ring strain could be
completely overturned in this catalyst-controlled
process. With these results, we propose that ge-
netically tunable enzymatic catalysis may provide
a general strategy to address the challenge of site
selectivity in C–H functionalization.
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