QM/MM study of the [4Fe-4S]-dependent (R)-2-hydroxyisocaproyl-CoA dehydratase: Dehydration via a redox pathway with an α-carbonyl radical intermediate

• Published in: Journal of catalysis 2023-05, Vol.421, p.419-430
• Main Authors: Wei, Wen-Jie, Liao, Rong-Zhen
• Format: Article
• Language: English
• Subjects: [4Fe-4S] cluster; catalytic activity; cis-trans isomers; Dehydratase; electron transfer; enzymes; family; fermentation; heterolytic cleavage; hydrogen bonding; leucine; ligands; oxygen; QM/MM calculations; Redox pathway; serine; Stereoselectivity
• ISSN: 0021-9517; 1090-2694
• DOI: 10.1016/j.jcat.2023.01.034

[Display omitted] •QM/MM approach was used to study the mechanism and stereoselectivity of the challenging β-hydrogen (pKa≈ 40) elimination reaction catalyzed by the [4Fe-4S]-dependent (R)-2-hydroxyisocaproyl-CoA dehydratase.•Different from the iron-sulfur containing radical S-adenosylmethionine (SAM) enzymes, the electron is directly transferred from the [4Fe-4S]1+ cluster to the substrate, and an α-carbonyl radical intermediate is formed with a spin-center shift process.•The calculations show that residues Ser37 and Glu55 play very important in the transfer of the electron from the [4Fe-4S]1+ cluster to the substrate.•By comparing all six possible antiferromagnetically-coupled doublet states, the calculations show that one alpha electron is transferred from the [4Fe-4S]1+ cluster to the substrate to achieve the maximum exchange and super exchange interactions, which can decrease the energy barriers.•The calculations in this work not only explain the current experimental results and can also be helpful for the understanding of the mechanisms and stereoselectivity of other members of the (R)-2-hydroxyacyl-CoA dehydratase enzyme family. The dehydration of proteinogenic amino acids is an essential fermentation process mediated by the (R)-2-hydroxyacyl-CoA dehydratase enzyme family. (R)-2-hydroxyisocaproyl-CoA dehydratase catalyzes the dehydration of (R)-2-hydroxyisocaproyl-CoA to harvest an E- product in the fermentation of L-leucine. In this work, QM/MM calculations were performed to elucidate the mechanism and stereoselectivity of this enzyme. Six possible antiferromagnetically coupled spin states (ααββ, αβαβ, αββα, ββαα, βαβα, βααβ) were considered for the [4Fe-4S] cluster. First, the binding mode of the substrate to the reduced [4Fe-4S]+ cluster switches from carbonyl oxygen coordination to α-hydroxy group coordination. This ligand exchange is coupled with the formation of a hydrogen bond between the substrate carbonyl oxygen and a second-shell serine residue, leading to an electron transfer from the [4Fe-4S]+ cluster to the substrate and the formation of a ketyl radical anion intermediate. This is followed by the heterolytic cleavage of the substrate C-OH bond, which is coupled with a spin-center shift process, generating a Fe-coordinated hydroxide and an α-carbonyl radical intermediate. Subsequently, the Fe-coordinated hydroxide abstracts the β-proton from the substrate, generating the experimentally identified allylic ketyl radical product. The syn-el