Abstract

Frequent occurrences of multi-drug resistance of pathogenic Gram-negative bacteria threaten human beings. The CMP-2-keto-3-deoxy-d-manno-octulosonic acid biosynthesis pathway is one of the new targets for antibiotic design. 2-Keto-3-deoxy-d-manno-octulosonate cytidylyltransferase (KdsB) is the key enzyme in this pathway. KdsB proteins from Burkholderia pseudomallei (Bp), B. thailandensis (Bt), Pseudomonas aeruginosa (Pa), and Chlamydia psittaci (Cp) have been assayed to find inhibitors. Interestingly, Rose Bengal (4,5,6,7-tetrachloro-2′,4′,5′,7′-tetraiodofluorescein) was turned out to be an inhibitor of three KdsBs (BpKdsB, BtKdsB, and PaKdsB) with promising IC₅₀ values and increased thermostability. The inhibitory enzyme kinetics of Rose Bengal revealed that it is competitive with 2-keto-3-deoxy-manno-octulosonic acid (KDO) but non-competitive against cytidine 5′-triphosphate (CTP). Induced-fit docking analysis of PaKdsB revealed that Arg160 and Arg185 together with other interactions in the substrate binding site seemed to play an important role in binding with Rose Bengal. We suggest that Rose Bengal can be used as the scaffold to develop potential antibiotics.

摘要

致病性革兰氏阴性细菌多药耐药性的频繁发生威胁着人类。的CMP-2-酮-3-脱氧- d -甘露-octulosonic酸生物合成途径是对抗生素设计新的目标之一。2-Keto-3- deoxy - d - manno - octulosonate cydydylyltransferase（KdsB）是该途径中的关键酶。来自假伯克霍尔德氏菌（Bp），泰国芽孢杆菌（Bt），铜绿假单胞菌（Pa）和鹦鹉热衣原体（Cp）的KdsB蛋白）已经过检测以找到抑制剂。有趣的是，玫瑰红（4,5,6,7-四氯-2'，4'，5'，7'-四碘荧光素）被证明是三种KdsB（Bp KdsB，Bt KdsB和Pa KdsB）的抑制剂。具有令人满意的IC ₅₀值和更高的热稳定性。玫瑰红的抑制酶动力学表明，它与2-酮-3-脱氧甘露糖辛酸（KDO）有竞争性，但与5'-三磷酸胞苷（CTP）没有竞争性。Pa的诱导拟合对接分析KdsB揭示Arg160和Arg185以及底物结合位点中的其他相互作用似乎在与Rose Bengal结合中起重要作用。我们建议将玫瑰孟加拉用作开发潜在抗生素的支架。