6IXM

Crystal structure of the ketone reductase ChKRED20 from the genome of Chryseobacterium sp. CA49 complexed with NAD

Summary for 6IXM

• Entry DOI: 10.2210/pdb6ixm/pdb
• Descriptor: Short-chain dehydrogenase reductase, NICOTINAMIDE-ADENINE-DINUCLEOTIDE (3 entities in total)
• Functional Keywords: oxidoreductase
• Biological source: Chryseobacterium sp. CA49
• Total number of polymer chains: 4
• Total formula weight: 107025.46

Authors

Zhao, F.J.,Jin, Y.,Liu, Z.C.,Wang, G.G.,Wu, Z.L. (deposition date: 2018-12-11, release date: 2019-04-03, Last modification date: 2023-11-22)

Primary citation

Li, T.B.,Zhao, F.J.,Liu, Z.,Jin, Y.,Liu, Y.,Pei, X.Q.,Zhang, Z.G.,Wang, G.,Wu, Z.L.
Structure-guided engineering of ChKRED20 from Chryseobacterium sp. CA49 for asymmetric reduction of aryl ketoesters.
Enzyme Microb. Technol., 125:29-36, 2019

PubMed Abstract: ChKRED20 is a robust NADH-dependent ketoreductase identified from the genome of Chryseobacterium sp. CA49 that can use 2-propanol as the ultimate reducing agent. The wild-type can reduce over 100 g/l ketones for some pharmaceutical relevant substrates, exhibiting a remarkable potential for industrial application. In this work, to overcome the limitation of ChKRED20 to aryl ketoesters, we first refined the X-ray crystal structure of ChKRED20/NAD complex at a resolution of 1.6 Å, and then performed three rounds of iterative saturation mutagenesis at critical amino acid sites to reshape the active cavity of the enzyme. For methyl 2-oxo-2-phenylacetate and ethyl 3-oxo-3-phenylpropanoate, several gain-of-activity mutants were achieved, and for ethyl 2-oxo-4-phenylbutanoate, improved mutants were achieved with k/K increasing to 196-fold of the wild-type. All three substrates were completely reduced at 100 g/l loading catalyzed with selected ChKRED20 mutants, and deliver the corresponding chiral alcohols with >90% isolated yield and 97 - >99%ee.

PubMed: 30885322
DOI: 10.1016/j.enzmictec.2019.03.001

Experimental method

X-RAY DIFFRACTION (1.601 Å)