Applied Sciences, Vol. 13, Pages 4493: A Screening Model of Antibacterial Agents Based on Escherichia coli Cell-Division Protein

Applied Sciences, Vol. 13, Pages 4493: A Screening Model of Antibacterial Agents Based on Escherichia coli Cell-Division Protein

Applied Sciences doi: 10.3390/app13074493

Authors: Qiuyu Fan Jianwen Wu Bolin Xi Chunxiao Li Xiumin Wang Huanrong Li

Pathogenic Escherichia coli cannot be killed by most antibiotics (including colistin, a last-resort drug) due to the rapid development of antibiotic resistance. A highly conserved prokaryotic mitotic protein, filamenting temperature-sensitive protein Z (FtsZ) with GTPase activity, plays a key role in cell division and has become a promising target for screening novel antibacterial agents. In this study, the amplified ftsZ gene was inserted into cloning/expression vectors and recombinantly produced in E. coli; the recombinant FtsZ protein was purified by the Ni2+-NTA affinity column and then was used to screen for natural antibacterial agents. The results showed that the ftsZ gene with a size of 1170 bp was successfully amplified from E. coli and inserted into the pET-28a expression vector. After induction with 0.2 mM isopropyl β-D-1-thiogalactopyranoside (IPTG), FtsZ was expressed in E. coli BL21 as inclusion bodies. After purification, the recombinant FtsZ protein showed GTPase activity. The highest GTPase activity (0.998 nmol/mL/min) of FtsZ was observed at a GTP concentration of 1.25 mM. Several alkaloids were screened by a constructed model of FtsZ inhibitors. Sanguinarine chloride exhibited higher antibacterial activity against E. coli and Salmonella enteritidis (with minimum inhibitory concentrations (MICs) of 0.04–0.16 mg/mL and minimum bactericidal concentrations (MBCs) of 0.16–0.32 mg/mL) than tetrandrine (0.16–0.32 mg/mL) and berberine hydrochloride (0.32–0.64 mg/mL). Berberine hydrochloride prevented FtsZ polymerization in a concentration-dependent manner and bound to FtsZ protein by hydrogen bonding interaction. This study suggested that the FtsZ-based E. coli screening model could be exploited for the development of novel antibacterial agents for clinical applications.