含铋配合物与奈瑟氏菌铁结合蛋白的反应性质研究

摘要

Ferric-ion binding protein (Fbp) plays a key role for capturing Fe3+ in pathogenic bacteria. Here, BiNTA·2H2O was syntheisized by reacting nitrilotriacetatic acid (H3NTA) with Bi(NO3)3 and characterized by element analysis and NMR spectroscopy. Fbp of Neisseria gonorrhoeae was expressed in E. Coli, and isolated and purified. The dynamics of the reaction of BiNTA with apo-Fbp was determined by UV-Vis spectroscopy at different ratio of nB6NTAapo-Fbp.The reaction follows first-order mechanism and the rate constant is (0.175 ±0.064) min-1 (in 10 mmol·L-1 Hepes/HPO42- buffer at pH 7.4 and 310 K). The similar kinetics and rate constant were observed for the reaction of NH4BiCit with apo-Fbp. Bi3+ can bind to apo-Fbp at 1:1 molar ratio, and the binding constant lgK is (21.43±0.20) for Fbp-Bi-NTA and (16.03±0.03) for Fbp-Bi-Cit, respectively. These results indicate that the Feu transfer protein Fbp in pathogens could be the potential target for the bismuth antibacterial drugs.%铁结合蛋白(Fbp)是致病菌获取Fe3+的关键蛋白.本文采用氨三乙酸(H3NTA)和硝酸铋反应制备BiNTA·2H2O,并运用元素分析、NMR等手段进行表征.通过在大肠杆菌中克隆表达和分离纯化出奈瑟氏淋病双球菌的Fbp,测定不同计量比nSiNTA apo-Fbp下反应的紫外可见光谱,确定BiNTA与apo-Fbp的反应为一级反应,反应速率常数约为(0.175-0.064) min-1(10 mmol· L-1 Hepes/HPO42- pH 7.4缓冲溶液,310K).NH4BiCit与apo-Fbp的反应也为一级反应,反应速率与BiNTA相近.Bi3+ apo-Fbp的饱和结合计量比为1∶1,形成三元配合物Fbp-Bi-NTA的结合常数(lgK)为(21.43±0.20),形成Fbp-Bi-Cit的结合常数(lgK)为(16.03±0.03).实验结果表明,致病菌中运输Fe3+的蛋白铁结合蛋白可作为含铋抗菌药物的潜在靶分子.