靶向UPP合成酶的抗耐药性粪肠球菌药物筛选及其抗菌机制探究  

Screening of the inhibitor of vancomycin-resistant Enterococcus faecalis UPPS and antibacterial mechanism of action study

作　　者：李维佳 佟书娟[1] 尚卓 王佳颖[1] 林炜 Li Wei-jia;Tong Shu-juan;Shang Zhuo;Wang Jia-ying;Lin Wei(School of Medicine&Holistic Integrative Medicine,Nanjing University of Chinese Medicine,Nanjing 210023)

机构地区：[1]南京中医药大学医学院·整合医学学院,南京210023

出　　处：《中国抗生素杂志》2021年第12期1102-1108,共7页Chinese Journal of Antibiotics

基　　金：国家自然科学基金(No.81903526和No.81703806);江苏省自然科学基金(No.BK20190798)。

摘　　要：目的筛选耐万古霉素粪肠球菌UPP合成酶(EfaUPPS)的特异性抑制剂,以期获得作用于该靶点的抗耐万古霉素粪肠球菌药物先导化合物。方法克隆并表达耐万古霉素粪肠球菌UPP合成酶,建立酶活力测定方法,通过比较化合物的最小抑菌浓度(MIC)、半数抑制浓度(IC_(50))、解离常数(K_(d)),并利用分子对接等方法对化合物进行活性评价和抗菌机制探讨。结果从老药库中筛选出了甲状腺激素类似物替拉曲可(tiratricol),发现其对EfaUPPS酶活力有中等强度的抑制效果,IC_(50)值为25.13μmol/L,由MST测得的K_(d)值为166±6.8μmol/L,对耐药性粪肠球菌的MIC值为16μg/mL。米氏方程的双倒数分析证明tiratricol是EfaUPPS的竞争性抑制剂,分子对接结果显示该化合物可能结合在EfaUPPS的底物结合口袋入口处,阻碍底物进入活性中心。结论甲状腺激素类似物tiratricol对EfaUPPS有中等程度的抑制活性,对其抗菌机制的深入理解将为抗耐药菌新型抗生素的设计提供新的思路。Objective To screen a specific inhibitor of vancomycin-resistant Enterococcus faecalis undecaprenyl pyrophosphate synthase(EfaUPPS)for antibacterial lead compounds.Methods The UPP synthase gene of vancomycin-resistant Enterococcus faecalis was cloned and expressed,the enzyme activity assay for EfaUPPS was established,and the inhibitory activity and preliminary molecular mechanism of compounds toward EfaUPPS were evaluated using minimum inhibitory concentration(MIC),half maximal inhibitory concentration(IC_(50)),and dissociation constant(K_(d))as well as molecular docking study.Results One EfaUPPS inhibitor candidate tiratricol from the old drug library showed medium inhibition to EfaUPPS with an IC_(50) of 25.13μmol/L,binding affinity to EfaUPPS with a K_(d) of(166±6.8)μmol/L,and antibacterial acitivty against vancomycin-resistant Enterococcus faecalis with a MIC of 16μg/mL.In addition,Lineweaver-Burk plots proves that tiratricol is a competitive inhibitor of EfaUPPS,molecular docking results revealed that tiratricol may bind to the entrance of the substrate binding pocket of EfaUPPS,and thus prevent the substrate from entering the active center of the enzyme.Conclusion Our results reported that the thyroid hormone analogue tiratricol can target the EfaUPPS enzyme and have moderate antibacterial activity against vancomycin-resistant Enterococcus faecalis for the first time.Further understanding of its antibacterial mechanism will provide new insight into the design of new antibiotics to combat drug-resistant bacterial infections.

关 键 词：耐万古霉素粪肠球菌 UPP合成酶抑制剂 替拉曲可 

分 类 号：R978.1[医药卫生—药品]