Novel Beta-Tubulin-Immobilized Nanoparticles Affinity Material for Screening beta-Tubulin Inhibitors from a Complex Mixture
Novel Beta-Tubulin-Immobilized Nanoparticles Affinity Material for Screening beta-Tubulin Inhibitors from a Complex Mixture
复制标题
用于从复杂混合物中筛选 β-微管蛋白抑制剂的新型 β-微管蛋白固定纳米颗粒亲和材料
DOI：
10.1021/acsami.6b13477
复制
发表时间：
2017
影响因子：
9.5
通讯作者：
Qin Xiaoyan
中科院分区：
文献类型：
--
作者：
Liu Qing-Shan;Deng Ran;Yang Qing-Fang;Cheng Lin;Luo Yongming;Li Keqin;Yin Xiaoying;Qin Xiaoyan
为了高效筛选和分离β-微管蛋白抑制剂，将β-微管蛋白固定在核壳结构的聚甲基丙烯酸甲酯/壳聚糖（PMMA/CS）纳米粒子上，制备了一种新型的固定化亲和材料--β-微管蛋白固定化纳米粒子（β-TIN）。以不同的对照药物对β-TIN的选择性和吸附性能进行了表征。比较了β-TIN、紫杉醇分子印迹聚合物（MIP）和C18吸附材料的选择性和富集率。采用β-TIN法从典型中药多穗金粟兰中筛选并分离了微管靶向抗肿瘤化合物。成功地从多穗金粟兰提取物中捕获了莪术醇、莪术呋喃和党参内酯三种活性成分。微量热泳实验表明，这三种化合物与β-微管蛋白有较强的结合作用，其解离常数（Kd）分别为1820 ± 0.68 nM、1640 ± 0.52 nM和284 ± 1.00 nM。此外，codonolactone与β-tubulin之间的结合亲和力大于紫杉醇与β-tubulin之间的结合亲和力。通过微管抑制模型证实了三个化合物的抗肿瘤活性，结果显示了与紫杉醇相似的抗肿瘤机制。通过分子动力学模拟，初步探讨了3种活性分子与β-微管蛋白的潜在结合位点及其构效关系。我们的研究首次报道了这种新型材料的使用，这种材料可以高效地从复杂的混合物中捕获低含量的β-微管蛋白抑制剂。筛选出的3个化合物均具有潜在的抗肿瘤活性，这些先导化合物具有新的作用机制，具有良好的开发前景。由于β-TIN易于制备，对靶蛋白具有良好的吸附性和选择性，并能有效地保持靶蛋白的空间构象和活性，因此在药物靶蛋白先导化合物的筛选中具有重要的应用价值。
In order to efficiently screen and isolate β-tubulin inhibitors, β-tubulin was immobilized on core–shell PMMA/CS (poly(methyl methacrylate)/Chitosan) nanoparticles to produce a new type of immobilized affinity material named β-tubulin-immobilized nanoparticles (β-TIN). The selectivity and adsorption performance of β-TIN were characterized using various control drugs. The β-TIN, the paclitaxel molecularly imprinted ploymers (MIP), and the C18 adsorbing material were compared for selectivity and enrichment ratio. Microtubule-targeting antitumor compounds were screened and isolated from a typical Chinese medicine,Chloranthus multistachys, by β-TIN. Three active compounds (curcolnol, zedoarofuran, and codonolactone) inChloranthus multistachysextract were captured successfully. Microscale thermophoresis demonstrated that these three compounds strongly bind to β-tubulin, and the dissociation constants (Kd) between the three active compounds and β-tubulin were 1820 ± 0.68 nM, 1640 ± 0.52 nM, and 284 ± 1.00 nM, respectively. Moreover, the binding affinity between codonolactone and β-tubulin was greater than that between paclitaxel and β-tubulin. The antitumor activities of the three compounds were confirmed by the microtubule inhibition model, and the results showed a similar antitumor mechanism as paclitaxel. Molecular dynamics simulations were performed to preliminarily investigate the potential binding sites and the structure–activity relationship between the three active molecules and β-tubulin. Our study is the first to report the use of this novel material which is highly efficient in capturing low-content β-tubulin inhibitors from a complex mixture. The three screened compounds exhibited potential antineoplastic activity, and these lead compounds utilize a new mechanism of action with promising development prospects. Because β-TIN is easily prepared, displays excellent adsorption and selectivity for targets, and can effectively maintain the steric conformation and activities of target proteins, it will be very useful in the screening of lead compounds for different drug target proteins.