Zhao Feitao, Fan Lili, Yang Jumei, Yang Mingjun, Zhang Chun, Wang Fang, Wang Yonggang
School of life science and Engineering, Lanzhou University of Technology, Lanzhou 730050, China.
Lanzhou University Second Hospital, Lanzhou 730000, China.
Int J Biol Macromol. 2024 Oct;277(Pt 2):133440. doi: 10.1016/j.ijbiomac.2024.133440. Epub 2024 Jun 28.
BACE1, a crucial enzyme in the amyloid-β deposition theory of Alzheimer's disease (AD), is targeted by Codonopsis pilosula, a traditional tonic believed to impede AD onset. However, the specific active compounds responsible for its effects remain elusive. Our prior network pharmacology research identified C. pilosula polysaccharides (CPPS) and Lobetyolin may serve as potential inhibitors of AD by suppressing amyloidogenesis. Here, we recombinantly expressed BACE1 under varied conditions and assessed its activity using Fluorescence Resonance Energy Transfer technology. Through spectroscopy, molecular docking, and dynamics, we elucidated the interactions of CPPS, Lobetyolin, and BACE1. Optimal BACE1 expression occurred at 22 °C with 0.4 mM IPTG for 6 h, yielding a 72 kDa protein. Enzyme kinetics displayed a maximum rate of 4096 μmol/min and a Michaelis constant of 16 mg/mL for BACE1. Spectroscopic analysis revealed differing binding affinities of the compounds at various temperatures, peaking at 293 K. Lobetyolin exhibited superior binding to BACE1 compared to CPPS, driven by hydrophobic and electrostatic forces. Molecular docking and dynamics highlighted hydrophobic amino acids' role in BACE1 interactions with Lobetyolin and CPPS, with binding energy < -1.2 kcal/mol signifying strong affinities. Notably, Lobetyolin and CPPS showed higher BACE1 affinity than APP, with the Lobetyolin-BACE1 complex being the most stable.
β-分泌酶1（BACE1）是阿尔茨海默病（AD）淀粉样蛋白β沉积理论中的一种关键酶，党参作为一种传统滋补品，被认为可以延缓AD的发病，它是BACE1的作用靶点。然而，其作用的具体活性化合物仍不清楚。我们之前的网络药理学研究表明，党参多糖（CPPS）和党参炔苷可能通过抑制淀粉样蛋白生成而成为AD的潜在抑制剂。在此，我们在不同条件下重组表达了BACE1，并使用荧光共振能量转移技术评估其活性。通过光谱学、分子对接和动力学研究，我们阐明了CPPS、党参炔苷和BACE1之间的相互作用。BACE1在22°C、0.4 mM异丙基硫代半乳糖苷（IPTG）诱导6小时的条件下表达最佳，产生一种72 kDa的蛋白质。酶动力学显示，BACE1的最大反应速率为4096 μmol/min，米氏常数为16 mg/mL。光谱分析表明，这些化合物在不同温度下具有不同的结合亲和力，在293 K时达到峰值。与CPPS相比，党参炔苷与BACE1的结合能力更强，这是由疏水作用和静电作用驱动的。分子对接和动力学研究突出了疏水氨基酸在BACE1与党参炔苷和CPPS相互作用中的作用，结合能 < -1.2 kcal/mol表示具有很强的亲和力。值得注意的是，党参炔苷和CPPS对BACE1的亲和力高于淀粉样前体蛋白（APP），其中党参炔苷 - BACE1复合物最稳定。
