Structure-guided selection of puromycin N-acetyltransferase mutants with enhanced selection stringency for deriving mammalian cell lines expressing recombinant proteins.

A clear trend was established; PAC mutants with low levels of enzyme activity such as Y171F generated very few colonies at low (1 μg/mL) puromycin concentrations and none at higher concentrations (3–5 μg/mL; Fig. 3 b), while PAC mutants with higher levels of enzyme activity such as Y30F, retained their ability to promote efficient colony formation across all puromycin concentrations used, albeit with fewer colonies than the WT (Fig. 3 b and Supplementary Table S4 ).

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