8JPT

Crystal Structure of the acyltransferase domain from the eighth module of the spinosad polyketide synthase

8JPT の概要

• エントリーDOI: 10.2210/pdb8jpt/pdb
• 分子名称: Polyene macrolide polyketide synthase (2 entities in total)
• 機能のキーワード: acyltransferase domain, transferase
• 由来する生物種: Saccharopolyspora spinosa
• タンパク質・核酸の鎖数: 3
• 化学式量合計: 132344.02

構造登録者

Huang, S.,Zheng, J. (登録日: 2023-06-12, 公開日: 2024-06-19, 最終更新日: 2025-07-02)

主引用文献

Huang, S.,Ji, H.,Zheng, J.
Structural and computational insights into the substrate specificity of acyltransferase domains from modular polyketide synthases.
Febs J., 291:3839-3855, 2024

PubMed Abstract: Polyketides are natural products synthesized by polyketide synthases (PKSs), where acyltransferase (AT) domains play a crucial role in selection of extender units. Engineering of AT domains enables the site-specific incorporation of non-natural extender units, leading to generation of novel derivatives. Here, we determined the crystal structures of AT domains from the fifth module of tylosin PKS (TylAT5) derived from Streptomyces fradiae and the eighth module of spinosad PKS (SpnAT8) derived from Saccharopolyspora spinosa, and combined them with molecular dynamics simulations and enzyme kinetic studies to elucidate the molecular basis of substrate selection. The ethylmalonyl-CoA-specific conserved motif TAGH of TylAT5 and the MMCoA-specific conserved motif YASH of SpnAT8 were identified within the substrate-binding pocket, and several key residues close to the substrate acyl moiety were located. Through site-directed mutagenesis of four residues near the active site, we successfully reprogrammed the specificity of these two AT domains toward malonyl-CoA. Mutations in TylAT5 enhanced its catalytic activity 2.6-fold toward malonyl-CoA, and mutations in SpnAT8 eliminated the substrate promiscuity. These results extend our understanding of AT substrate specificity and would benefit the engineering of PKSs.

PubMed: 38922792
DOI: 10.1111/febs.17206

実験手法

X-RAY DIFFRACTION (3.26 Å)