Abstract：
l-(+)-Tartaric acid plays important roles in various industries, including pharmaceuticals, foods, and chemicals. cis-Epoxysuccinate hydrolases (CESHs) are crucial for converting cis-epoxysuccinate to l-(+)-tartrate in the industrial production process. There is, however, a lack of detailed structural and mechanistic information on CESHs, limiting the discovery and engineering of these industrially relevant enzymes. In this study, we report the crystal structures of RoCESH and KoCESH-l-(+)-tartrate complex. These structures reveal the key amino acids of the active pocket and the catalytic triad residues and elucidate a dynamic catalytic process involving conformational changes of the active site. Leveraging the structural insights, we identified a robust BmCESH (550 ± 20 U·mg-1) with sustained catalytic activity even at a 3 M substrate concentration. After six batches of transformation, immobilized cells with overexpressed BmCESH maintained 69% of their initial activity, affording an overall productivity of 200 g/L/h. These results provide valuable insights into the development of high-efficiency CESHs and the optimization of biotransformation processes for industrial uses.
摘要：
l-(+)-酒石酸在各行各业中发挥着重要作用,包括药品,食物,和化学品。顺式-环氧琥珀酸水解酶（CESH）对于在工业生产过程中将顺式-环氧琥珀酸转化为1-（）-酒石酸盐至关重要。有,然而,缺乏关于CESH的详细结构和机械信息，限制了这些工业相关酶的发现和工程。在这项研究中,我们报道了RoCESH和KoCESH-l-(+)-酒石酸盐络合物的晶体结构。这些结构揭示了活性口袋和催化三联体残基的关键氨基酸，并阐明了涉及活性位点构象变化的动态催化过程。利用结构性见解，我们确定了一个强大的BmCESH（550±20U·mg-1），即使在3M底物浓度下也具有持续的催化活性。经过六批改造,具有过表达BmCESH的固定化细胞保持其初始活性的69％，提供200克/升/小时的总生产率。这些结果为开发高效CESH和优化工业用途的生物转化过程提供了有价值的见解。
