Industrial Enzymes National Engineering Laboratory, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China.
Org Biomol Chem. 2019 Feb 20;17(8):2070-2076. doi: 10.1039/c8ob02758g.
LepI is a novel multifunctional enzyme that catalyzes stereoselective dehydration, Diels-Alder reaction, and retro-Claisen rearrangement. Here we report the crystal structure of LepI in complex with its co-factor S-adenosyl methionine (SAM). LepI forms a tetramer via the N-terminal helical domain and binds to a SAM molecule in the C-terminal catalytic domain. The binding modes of various LepI substrates are investigated by docking simulations, which suggest that the substrates are bound via both hydrophobic and hydrophilic forces, as well as cation-π interactions with the positively charged SAM. The reaction starts with a dehydration step in which H133 possibly deprotonates the pyridone hydroxyl group of the substrate, while D296 might protonate an alkyl-chain hydroxyl group. Subsequent pericyclization may be facilitated by the correct fold of the substrate's alkyl chain and a thermodynamic driving force towards σ-bonds at the expense of π-bonds. These results provide structural insights into LepI catalysis and are important in understanding the mechanism of enzymatic pericyclization.
LepI 是一种新型多功能酶，可催化立体选择性脱水、Diels-Alder 反应和 retro-Claisen 重排。在这里，我们报告了 LepI 与其辅因子 S-腺苷甲硫氨酸（SAM）复合物的晶体结构。LepI 通过 N 端螺旋结构域形成四聚体，并结合到 C 端催化结构域中的一个 SAM 分子上。通过对接模拟研究了各种 LepI 底物的结合模式，表明底物通过疏水和亲水相互作用以及与带正电荷的 SAM 的阳离子-π 相互作用结合。反应首先通过脱水步骤开始，其中 H133 可能使底物吡啶酮羟基去质子化，而 D296 可能使烷基链羟基质子化。随后的周环化可能得益于底物烷基链的正确折叠和朝着 σ 键而不是 π 键的热力学驱动力。这些结果提供了 LepI 催化的结构见解，对于理解酶促周环化的机制非常重要。
