8J85

Cryo-EM structure of ochratoxin A-detoxifying amidohydrolase ADH3 mutant S88E in complex with ochratoxin A

8J85 の概要

• エントリーDOI: 10.2210/pdb8j85/pdb
• EMDBエントリー: 36060
• 分子名称: Amidohydrolase family protein, ZINC ION, (2~{S})-2-[[(3~{R})-5-chloranyl-3-methyl-8-oxidanyl-1-oxidanylidene-3,4-dihydroisochromen-7-yl]carbonylamino]-3-phenyl-propanoic acid (3 entities in total)
• 機能のキーワード: amidohydrolase, octamer, ochratoxin a degradtion, cryo-em structure, hydrolase
• 由来する生物種: Stenotrophomonas acidaminiphila
• タンパク質・核酸の鎖数: 8
• 化学式量合計: 370346.39

構造登録者

Dai, L.H.,Niu, D.,Huang, J.-W.,Li, X.,Shen, P.P.,Li, H.,Hu, Y.M.,Yang, Y.,Chen, C.-C.,Guo, R.-T. (登録日: 2023-04-30, 公開日: 2023-08-30, 最終更新日: 2025-07-02)

主引用文献

Dai, L.,Niu, D.,Huang, J.W.,Li, X.,Shen, P.,Li, H.,Xie, Z.,Min, J.,Hu, Y.,Yang, Y.,Guo, R.T.,Chen, C.C.
Cryo-EM structure and rational engineering of a superefficient ochratoxin A-detoxifying amidohydrolase.
J Hazard Mater, 458:131836-131836, 2023

PubMed Abstract: Ochratoxin A (OTA) is among the most prevalent mycotoxins detected in agroproducts, posing serious threats to human and livestock health. Using enzymes to conduct OTA detoxification is an appealing potential strategy. The recently identified amidohydrolase from Stenotrophomonas acidaminiphila, termed ADH3, is the most efficient OTA-detoxifying enzyme reported thus far and can hydrolyze OTA to nontoxic ochratoxin α (OTα) and L-β-phenylalanine (Phe). To elucidate the catalytic mechanism of ADH3, we solved the single-particle cryo-electron microscopy (cryo-EM) structures of apo-form, Phe- and OTA-bound ADH3 to an overall resolution of 2.5-2.7 Å. The role of OTA-binding residues was investigated by structural, mutagenesis and biochemical analyses. We also rationally engineered ADH3 and obtained variant S88E, whose catalytic activity was elevated by 3.7-fold. Structural analysis of variant S88E indicates that the E88 side chain provides additional hydrogen bond interactions to the OTα moiety. Furthermore, the OTA-hydrolytic activity of variant S88E expressed in Pichia pastoris is comparable to that of Escherichia coli-expressed enzyme, revealing the feasibility of employing the industrial yeast strain to produce ADH3 and its variants for further applications. These results unveil a wealth of information about the catalytic mechanism of ADH3-mediated OTA degradation and provide a blueprint for rational engineering of high-efficiency OTA-detoxifying machineries.

PubMed: 37331057
DOI: 10.1016/j.jhazmat.2023.131836

実験手法

ELECTRON MICROSCOPY (2.7 Å)