Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, Mexico City, Mexico.
Instituto de Investigaciones Biomédicas, Universidad Nacional Autónoma de México, Mexico City, Mexico
J Bacteriol. 2019 Aug 8;201(17). doi: 10.1128/JB.00177-19. Print 2019 Sep 1.
OmpA-like proteins are involved in the stabilization of the outer membrane, resistance to osmotic stress, and pathogenesis. In , OmpA2 forms a physiologically relevant concentration gradient that forms by an uncharacterized mechanism, in which the gradient orientation depends on the position of the gene locus. This suggests that OmpA2 is synthesized and translocated to the periplasm close to the position of the gene and that the gradient forms by diffusion of the protein from this point. To further understand how the OmpA2 gradient is established, we determined the localization and mobility of the full protein and of its two structural domains. We show that OmpA2 does not diffuse and that both domains are required for gradient formation. The C-terminal domain binds tightly to the cell wall and the immobility of the full protein depends on the binding of this domain to the peptidoglycan; in contrast, the N-terminal membrane β-barrel diffuses slowly. Our results support a model in which once OmpA2 is translocated to the periplasm, the N-terminal membrane β-barrel is required for an initial fast restriction of diffusion until the position of the protein is stabilized by the binding of the C-terminal domain to the cell wall. The implications of these results on outer membrane protein diffusion and organization are discussed. Protein concentration gradients play a relevant role in the organization of the bacterial cell. The protein OmpA2 forms an outer membrane polar concentration gradient. To understand the molecular mechanism that determines the formation of this gradient, we characterized the mobility and localization of the full protein and of its two structural domains an integral outer membrane β-barrel and a periplasmic peptidoglycan binding domain. Each domain has a different role in the formation of the OmpA2 gradient, which occurs in two steps. We also show that the OmpA2 outer membrane β-barrel can diffuse, which is in contrast to what has been reported previously for several integral outer membrane proteins in , suggesting a different organization of the outer membrane proteins.
OmpA 样蛋白参与稳定外膜、抵抗渗透压应激和发病机制。在 中，OmpA2 形成一种生理相关的浓度梯度，其形成机制尚不清楚，梯度方向取决于基因座的位置。这表明 OmpA2 是在靠近基因位置的周质中合成和转运的，并且梯度是通过蛋白质从该点扩散形成的。为了进一步了解 OmpA2 梯度是如何建立的，我们确定了完整蛋白及其两个结构域的定位和迁移率。我们表明 OmpA2 不会扩散，并且梯度形成需要两个结构域。C 端结构域与细胞壁紧密结合，完整蛋白的不流动性取决于该结构域与肽聚糖的结合；相比之下，N 端膜 β-桶缓慢扩散。我们的结果支持这样一种模型，即一旦 OmpA2 转运到周质中，N 端膜 β-桶就需要在蛋白质的位置通过与细胞壁结合而稳定之前，初始快速限制扩散。这些结果对外膜蛋白扩散和组织的影响进行了讨论。蛋白质浓度梯度在外膜蛋白扩散和组织中起着重要作用。OmpA2 形成一种外膜极性浓度梯度。为了了解决定该梯度形成的分子机制，我们对完整蛋白及其两个结构域的迁移率和定位进行了表征，这两个结构域分别是一个完整的外膜β-桶和一个周质肽聚糖结合域。每个结构域在外膜蛋白形成梯度中都有不同的作用，该梯度分两步形成。我们还表明，OmpA2 外膜 β-桶可以扩散，这与之前在 中报道的几种完整外膜蛋白的情况形成对比，这表明外膜蛋白的组织方式不同。
