The Molecular Basis Of Antibiotic Resistance

大肠杆菌的marRAB多重抗生素抗性操纵子通过复杂的反应网络控制大量基因的表达，导致低水平的抗生素和超氧化物抗性。 马尔R自动抑制mar操纵子，但在与水杨酸相互作用后失活，失去其DNA结合能力。 这又导致操纵子的去阻遏和MarA的表达，其激活约40至60个启动子（mar/sox/rob调节子）的转录，包括marRAB启动子本身（自激活）。 我们最近集中在MarA激活这些不同启动子的能力上，试图建立一个数学模型来描述网络。 我们发现，MarA和/或SoxS的浓度需要导致半最大刺激变化至少30倍的不同的启动子，是相似的，但不相同的两个激活剂。因此，在激活剂的中间浓度下，虽然调节子中的一些基因表现出显著的激活，但其他基因保持休眠。 我们把这种类型的调节称为分层激活。一个数学模型已被开发来定量描述这种现象。该模型的显著结论是，调节子的大多数基因的激活最好由正向反应速率（清除率）的非常大的增加来解释，伴随着在激活剂存在下RNA聚合酶对启动子的亲和力的降低（而不是增加）。我们和其他人以前提出了模型的相互作用的MarA与RNA聚合酶的转录激活所必需的。 上述分析使人们对这些模式的某些方面产生了怀疑。因此，我们目前正在使用上述技术对MarA突变体进行广泛的分析。此外，我们还对两个启动子进行了广泛的分析，这两个启动子被认为调节主要负责抗生素抗性的三个基因tolC和acrAB。 这项工作主要是与J.L.博士合作进行的。Rosner和Michael Wall（计算机和计算科学与生物科学，Los Alamos国家实验室，Mail Stop B256，Los Alamos NM 87545 USA）。

The marRAB multiple antibiotic resistance operon of Escherichia coli controls the expression of a large number of genes resulting in low level antibiotic and superoxide resistance through a complex network of reactions. MarR auto-represses the mar operon but is inactivated upon interaction with salicylate, losing its DNA binding capacity. This, in turn, results in derepression of the operon and expression of MarA, which activates the transcription of some 40 to 60 promoters (the mar/sox/rob regulon) including the marRAB promoter itself (auto-activation). We have focused recently on the capacity of MarA to activate these disparate promoters in an attempt to develop a mathematical model to describe the network. We find that the concentration of MarA and/or SoxS necessary to result in half-maximum stimulation varies by at least 30-fold for the different promoters and is similar but not identical for the two activators. Thus, at intermediate concentrations of the activators, while some genes in the regulon exhibit significant activation, others remain dormant. We refer to this type of regulation as hierarchical activation. A mathematical model has been developed to characterize this phenomenon quantitatively. The remarkable conclusion of this modeling is that the activation of most genes of the regulon is best accounted for by a very large increase in the forward reaction rate (clearance) accompanied by a decrease (rather than an increase) in the affinity of RNA polymerase for the promoter in the presence of the activator. We and others have previously proposed models for the interaction of MarA with RNA polymerase requisite for transcriptional activation. The above analyses have thrown into doubt some aspects of those models. Consequently we are currently engaged in an extensive analysis of MarA mutants employing the above technique. In addition we have carried out an extensive analysis of the two promoters believed to regulate the three genes primarily responsible for antibiotic resistance, tolC and acrAB. This work was carried out in collaboration principally with Drs. J.L. Rosner and Michael Wall (Computer and Computational Sciences & Bioscience, Los Alamos National Laboratory, Mail Stop B256,Los Alamos NM 87545 USA).