Zhou_2022_ACS.Catal_12_762

Reference

Title : Structural and Mechanistic Insights into Chain Release of the Polyene PKS Thioesterase Domain - Zhou_2022_ACS.Catal_12_762
Author(s) : Zhou Y , Tao W , Qi Z , Wei J , Shi T , Kang Q , Zheng J , Zhao Y , Bai L
Ref : ACS Catal , 12 :762 , 2022
Abstract :

Polyketides serve as rich source of therapeutically relevant drug leads. The manipulation of polyketide synthases (PKSs) for generating derivatives with improved activities usually results in substantially reduced yields. Growing evidence suggests that type I PKS thioesterase (TE) domains are key bottlenecks in the biosynthesis of polyene antibiotics, such as pimaricin and amphotericin, and their unnatural derivatives. Herein, we elucidate the structure of the 26-membered macrolide-complexed TE domain from the pimaricin pathway (Pim TE), which specifies a spacious bifunnel-shaped substrate channel with a highly hydrophobic cleft proximal to the catalytic triad and a hydrophilic loop I region specific for the cyclization of amphiphilic polyene macrolide. Notably, the natural intermediate with C12-COOH is stabilized by a hydrogen-bond network, as well as by interactions between the polyene moiety and the hydrophobic cleft. Moreover, the bottleneck in processing the unnatural intermediate with C12-CH3 is attributed to the unstable and mismatched docking of the curved substrate in the channel. Aided by an in vitro assay with a fully elongated linear polyene intermediate as the substrate, multiple strategies were adopted, herein, to engineer Pim TE, including introducing H-bond donors, enhancing hydrophobic interactions, and modifying the catalytic center. Efficient TE mutations with increased substrate conversion up to 39.2% in vitro were further conducted in vivo, with a titer increase as high as 37.1% for the less toxic decarboxylated pimaricin derivatives with C12-CH3. Our work uncovers the mechanism of TE-catalyzed polyene macrolide formation and highlights TE domains as targets for PKS manipulation for titer increases in engineered unnatural polyketide derivatives.

PubMedSearch : Zhou_2022_ACS.Catal_12_762
PubMedID:
Gene_locus related to this paper: 9actn-f1cla7

Related information

Inhibitor 4d-81E-7VO5
Substrate Pimaricin-linear-analogue-synthetic    Pimaricin
Gene_locus 9actn-f1cla7
Family Thioesterase
Structure 7VO4    7VO5

Citations formats

Zhou Y, Tao W, Qi Z, Wei J, Shi T, Kang Q, Zheng J, Zhao Y, Bai L (2022)
Structural and Mechanistic Insights into Chain Release of the Polyene PKS Thioesterase Domain
ACS Catal 12 :762

Zhou Y, Tao W, Qi Z, Wei J, Shi T, Kang Q, Zheng J, Zhao Y, Bai L (2022)
ACS Catal 12 :762