P450 fatty acid
decarboxylases (FADCs) have recently been attracting considerable attention owing
to their one-step direct production of industrially important 1-alkenes from biologically abundant
feedstock free fatty acids under mild conditions. However, attempts to improve
the catalytic activity of FADCs have met with little success. Protein
engineering has been limited to selected residues and small mutant libraries
due to lack of an effective high-throughput screening (HTS) method. Here, we
devise a catalase-deficient <i>Escherichia
coli</i> host strain and report an HTS approach based on colorimetric detection
of H<sub>2</sub>O<sub>2</sub>-consumption activity of FADCs. Directed evolution
enabled by this method has led to effective identification for the first time
of improved FADC variants for medium-chain 1-alkene production from both DNA
shuffling and random mutagenesis libraries. Advantageously, this screening
method can be extended to other enzymes that stoichiometrically utilize H<sub>2</sub>O<sub>2</sub>
as co-substrate.
