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Publication Abstract from PubMed

The evolution of a promiscuous enzyme for its various activities often results in catalytically specialized variants. This is an important natural mechanism to ensure the proper functioning of natural metabolic networks. It also acts as both a curse and blessing for enzyme engineers, where enzymes that have undergone directed evolution may exhibit exquisite selectivity at the expense of a diminished overall catalytic repertoire. We previously performed two independent directed evolution campaigns on a promiscuous designer enzyme that leverages the unique properties of a noncanonical amino acid (ncAA) para-aminophenylalanine (pAF) as catalytic residue, resulting in two evolved variants which are both catalytically specialized. Here, we combine mutagenesis, crystallography, and computation to reveal the molecular basis of the specialization phenomenon. In one evolved variant, an unexpected change in quaternary structure biases substrate dynamics to promote enantioselective catalysis, while the other demonstrates synergistic cooperation between natural side chains and the pAF residue to form semisynthetic catalytic machinery.

Evolutionary Specialization of a Promiscuous Designer Enzyme.,Leveson-Gower RB, Tiessler-Sala L, Rozeboom HJ, Thunnissen AWH, Marechal JD, Roelfes G ACS Catal. 2025 Jan 13;15(3):1544-1552. doi: 10.1021/acscatal.4c06409. , eCollection 2025 Feb 7. PMID:39944761^[1]

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine.