ENGINEERING THE SUBSTRATE SCOPE OF THE FE(II) DEPENDENT HALOGENASE WELO15

摘要

Selective halogenation is an important reaction for late-stage functionalisation of drug-like molecules. Performing halogenations under mild conditions using sodium chloride as the chlorine source has great potential for sustainable catalysis. The discovery of non-heme iron (NHI) and 2-oxoglutarate dependent halogenases, acting directly on a small organic molecule and not on acyl-carrier bound substrates, has eliminated a major drawback of know NHI-halogenases. Hence, these enzymes represent attractive starting points for developing biocatalytic routs for selective, aliphatic chlorination, a paramount challenge in organic synthesis. The wild-types have a narrow natural substrate-scope and are unexplored for biocatalytic applications. After solving the crystal structure of WelO15 from Westiella intricata, we used directed evolution to redesign the active site using a small-but-smart amino acid alphabet, thereby limiting the screening effort to a HPLC compatible throughput. New variants were found, able to chlorinate novel synthesized non-natural substrates. This study represents a first step towards milder, selective chlorination using biocatalysis.