Molecular Interactions of Cephalosporins with the Deep Binding Pocket of the RND Transporter AcrB

The drug/proton antiporter AcrB, part of the major efflux pump AcrABZ-TolC in Escherichia coli, is characterized by its impressive ability to transport chemically diverse compounds, conferring a multidrug resistance phenotype. However, the molecular features differentiating between good and poor substrates of the pump have yet to be identified. In this work, we combined molecular docking with molecular dynamics simulations to study the interactions between AcrB and two representative cephalosporins, cefepime and ceftazidime (a good and poor substrate of AcrB, respectively). Our analysis revealed different binding preferences of the two compounds toward the subsites of the large deep binding pocket of AcrB. Cefepime, although less hydrophobic than ceftazidime, showed a higher affinity than ceftazidime for the so-called hydrophobic trap, a region known for binding inhibitors and substrates. This supports the hypothesis that surface complementarity between the molecule and AcrB, more than the intrinsic hydrophobicity of the antibiotic, is a feature required for the interaction within this region. Oppositely, the preference of ceftazidime for binding outside the hydrophobic trap might not be optimal for triggering allosteric conformational changes needed to the transporter to accomplish its function. Altogether, our findings could provide valuable information for the design of new antibiotics less susceptible to the efflux mechanism.大肠杆菌（Escherichia coli）主要外排泵AcrABZ-TolC的组成部分——药物/质子反向转运蛋白（drug/proton antiporter）AcrB，以其能够转运化学结构多样的化合物、赋予宿主多药耐药表型的卓越能力而广受关注。然而，区分该外排泵优质底物与劣质底物的分子特征至今仍未被阐明。本研究将分子对接（molecular docking）与分子动力学模拟（molecular dynamics simulations）相结合，针对AcrB与两种代表性头孢菌素类（cephalosporins）化合物——头孢吡肟（cefepime）与头孢他啶（ceftazidime，分别为AcrB的优质与劣质底物）的相互作用展开了系统研究。分析结果显示，两种化合物对AcrB庞大深邃的结合口袋的不同亚位点具有差异化结合偏好。尽管头孢吡肟的疏水性弱于头孢他啶，但其与所谓的疏水陷阱（hydrophobic trap）——一个已知可结合抑制剂与底物的区域——的亲和力却更高。这支持了如下假说：相较于抗生素自身的固有疏水性，分子与AcrB之间的表面互补性，才是该区域内相互作用所需的关键特征。与之相反，头孢他啶倾向于结合疏水陷阱以外的区域，这或许无法最优触发转运蛋白完成其功能所需的变构构象变化。综上，本研究结果可为设计更不易受外排机制影响的新型抗生素提供极具价值的参考依据。