Functional characterization and structural basis of an efficient ochratoxin A-degrading amidohydrolase
Ochratoxin A (OTA) contamination in various agro-products poses a serious threat to the global food safety and human health, leading to enormous economic losses. Enzyme-mediated OTA degradation is an appealing strategy, and the search for more efficient enzymes is a prerequisite for achieving this goal. Here, a novel amidohydrolase, termed Pw ADH, was demonstrated to exhibit 7.3-fold higher activity than that of the most efficient OTA-degrading ADH3 previously reported. Cryo-electron microscopy structure analysis indicated that additional hydrogen-bond interactions among OTA and the adjacent residue H163, the more compact substrate-binding pocket, and the wider entry to the substrate-access cavity might account for the more efficient OTA-degrading activity of Pw ADH compared with that of ADH3. We conducted a structure-guided rational design of Pw ADH and obtained an upgraded variant, G88D, whose OTA-degrading activity was elevated by 1.2-fold. In addition, Pw ADH and the upgraded G88D were successfully expressed in the industrial yeast Pichia pastoris , and their catalytic activities were compared to those of their counterparts produced in E. coli , revealing the feasibility of producing Pw ADH and its variants in industrial yeast strains. These results illustrate the structural basis of a novel, efficient OTA-degrading amidohydrolase and will be beneficial for the development of high-efficiency OTA-degrading approaches.
危险品化学品经营许可证（带存储）