Department of Biosciences, Rice University, Houston, TX, USA.
Department of Applied Mathematics & Statistics, University of California, Santa Cruz, CA, USA.
Mol Syst Biol. 2017 Dec 22;13(12):964. doi: 10.15252/msb.20177635.
The major facilitator superfamily (MFS) effluxers are prominent mediators of antimicrobial resistance. The biochemical characterization of MFS proteins is hindered by their complex membrane environment that makes biochemical analysis challenging. Since the physicochemical properties of proteins drive the fitness of an organism, we posed the question of whether we could reverse that relationship and derive meaningful biochemical parameters for a single protein simply from fitness changes it confers under varying strengths of selection. Here, we present a physiological model that uses cellular fitness as a proxy to predict the biochemical properties of the MFS tetracycline efflux pump, TetB, and a family of single amino acid variants. We determined two lumped biochemical parameters roughly describing and for TetB and variants. Including protein levels into our model allowed for more specified prediction of pump parameters relating to substrate binding affinity and pumping efficiency for TetB and variants. We further demonstrated the general utility of our model by solely using fitness to assay a library of variants and estimate their biochemical properties.
主要易化超家族（MFS）外排泵是抗菌药物耐药性的主要介导者。由于其复杂的膜环境，使得生化分析具有挑战性，因此 MFS 蛋白的生化特征难以确定。由于蛋白质的理化性质决定了生物体的适应性，我们提出了这样一个问题：我们是否可以逆转这种关系，并根据它在不同选择压力下赋予的适应性变化，为单个蛋白质推导出有意义的生化参数。在这里，我们提出了一个生理模型，该模型使用细胞适应性作为替代物来预测 MFS 四环素外排泵 TetB 和一系列单个氨基酸变体的生化特性。我们确定了两个大致描述 TetB 和变体的聚集生化参数。将 蛋白水平纳入我们的模型，使得可以更具体地预测 TetB 和变体与底物结合亲和力和泵送效率相关的泵参数。我们通过仅使用适应性来测定一个包含 个变体的文库并估计它们的生化特性，进一步证明了我们模型的通用性。
