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Re‐Programming and Optimization of a L‐Proline cis‐4‐Hydroxylase for the cis‐3‐Halogenation of its Native Substrate.
- Source :
-
ChemCatChem . 9/17/2021, Vol. 13 Issue 18, p3914-3919. 6p. - Publication Year :
- 2021
-
Abstract
- Non‐heme iron/α‐ketoglutarate dependent halogenases acting on freestanding substrates catalyze the regio‐ and stereoselective halogenation of inactivated C(sp3)‐H bonds. Yet, with only a handful of these halogenases characterized, the biosynthetic potential of enzymatic radical halogenation remains limited. Herein, we describe the remodeling of L‐proline cis‐4‐hydroxylase from Sinorhizobium meliloti into a halogenase by introduction of a single point mutation (D108G) into the enzyme's active site. The re‐programmed halogenase displays a striking regio‐divergent reaction chemistry: While halogenation of L‐proline exclusively occurs at the C3‐position, the retained hydroxylation activity leads to derivatization at the C4‐position, corresponding to the regioselectivity of the wildtype enzyme. By employing several rounds of directed evolution, an optimized halogenase variant with 98‐fold improved apparent kcat/Km for chlorination of L‐proline compared to the parental enzyme SmP4H (D108G) was identified. The development and optimization of this novel halogenation biocatalyst highlights the possibility to rationally harness the chemical versatility of non‐heme Fe/αKG dependent dioxygenases for C−H functionalization. [ABSTRACT FROM AUTHOR]
Details
- Language :
- English
- ISSN :
- 18673880
- Volume :
- 13
- Issue :
- 18
- Database :
- Academic Search Index
- Journal :
- ChemCatChem
- Publication Type :
- Academic Journal
- Accession number :
- 152513744
- Full Text :
- https://doi.org/10.1002/cctc.202100591