[Catalytic mechanism of pterocarpan reductase in Lotus corniculatus]

The roots of Lotus corniculatus, a Chinese medicinal herb, have been used in traditional Chinese medicine(TCM) to treat various diseases, including coughing due to wind-heat, fever due to Yin deficiency, eczema, and hemorrhoids. In recent years, pharmacological studies have revealed that the flavonoids in L. corniculatus, such as(-)-vestitol, exhibit antiviral, anticancer, and anti-inflammatory activities. However, the exploitation of its resources has been limited due to the low content in plants and the high difficulty of chemical synthesis. To guide the enhancement of the content of active substances such as(-)-vestitol in plants, this study investigated the catalytic mechanism of pterocarpan reductase(PTR) involved in the biosynthesis. Molecular docking and site-directed mutagenesis were employed to reveal the interaction modes between PTR and its substrates, which identified the crucial role of the lysine(Lys) residue in the catalytic process. The catalytic process of substrate is driven by the abstraction of the hydrogen atom of the hydroxyl group on the A-ring of the substrate, which leads to the formation of the transition state. Additionally, other amino acid residues, including isoleucine(Ile), phenylalanine(Phe), and tyrosine(Tyr), play crucial roles in the catalytic process through hydrophobic interactions, π-π stacking effects, and hydrogen bonding interactions, which provided a structural basis for understanding PTR catalytic characteristics. These findings not only give insights into the biosynthetic pathways of isoflavonoids in L. corniculatus but also offer a theoretical foundation for guiding the enhancement of flavonoid content in TCM, which are expected to promote the application and development of TCM in modern medicine.