Figure 4.
Genetic engineering of the ArM. (A) Analysis of amino acids for mutagenesis based on crystallography (PDB: 5vnp).34 Seven sites were targeted for saturation mutagenesis and activity profiling: E133, E143, F144, M175, P243, V245, and L271. Mutations at five of these sites increased the catalytic performance marginally. The best hits are displayed in panel B. The wt HT samples were completed as biological replicates, and the variants are replicates from the same protein purification batch. The bioconjugation and metathesis reactions were conducted as described for Figure 3.