Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Kongens Lyngby, Denmark.
Department of Biology and Biological Engineering, Chalmers University of Technology, Göteborg, 41296, Sweden.
Int J Nanomedicine. 2022 Dec 28;17:6707-6721. doi: 10.2147/IJN.S387590. eCollection 2022.
Antibacterial activity of graphene oxide (GO) has been extensively studied, wherein penetration of the bacterial cell membrane and oxidative stress are considered to play a major role in the bactericidal activity of GO. However, the specific mechanism responsible for the antibacterial activity of GO remains largely unknown. Hence, the goal of this study was to explore the mode of action of GO, via an in-depth proteomic analysis of the targeted bacteria.
was grown in the presence of GO and samples were collected at different growth phases to examine the cell viability and to analyze the changes in protein expression. Antimicrobial efficiency of GO was tested by assessing bacterial viability, live/dead staining and scanning electron microscopy. The intracellular reactive oxygen species (ROS) induced by GO treatment were examined by fluorescence microscopy. Label-free quantitative proteomics analysis was performed to examine the differentially regulated proteins in after GO treatment.
GO treatment was observed to reduce viability, from 50 ± 17% after 4 h, to 93 ± 2% after 24 h. The live/dead staining confirmed this progressive antimicrobial effect of GO. SEM images revealed the wrapping of bacterial cells and their morphological disruption by means of pore formation due to GO insertion. GO treatment was observed to generate intracellular ROS, correlating to the loss of cell viability. The proteomics analysis revealed alteration in the expression of cell membrane, oxidative stress response, general stress response, and virulence-associated proteins in GO-treated bacterial cells. The time-dependent bactericidal activity of GO correlated with a higher number of differentially regulated proteins involved in the above.-mentioned processes.
The obtained results suggest that the time-dependent bactericidal effect of GO is attributed to its wrapping/trapping ability, ROS production and due to physical disruption of the cell membrane.
氧化石墨烯（GO）的抗菌活性已得到广泛研究，其中细胞膜穿透和氧化应激被认为在 GO 的杀菌活性中起主要作用。然而，GO 抗菌活性的确切机制在很大程度上仍不清楚。因此，本研究的目的是通过对目标细菌进行深入的蛋白质组学分析来探索 GO 的作用方式。
在存在 GO 的情况下培养 ，并在不同的生长阶段收集样品，以检查细胞活力并分析蛋白质表达的变化。通过评估 GO 对细菌活力的影响、死活染色和扫描电子显微镜来测试 GO 的抗菌效率。通过荧光显微镜检查 GO 处理诱导的细胞内活性氧 (ROS)。通过无标记定量蛋白质组学分析检查 GO 处理后 中差异调节的蛋白质。
GO 处理被观察到降低了 的活力，从 4 小时后的 50 ± 17%降低到 24 小时后的 93 ± 2%。死活染色证实了 GO 的这种渐进式抗菌作用。SEM 图像显示，由于 GO 的插入，细菌细胞被包裹并通过形成孔导致其形态破坏。GO 处理被观察到产生细胞内 ROS，与细胞活力丧失相关。蛋白质组学分析显示，GO 处理的细菌细胞中细胞膜、氧化应激反应、一般应激反应和毒力相关蛋白的表达发生改变。GO 的时间依赖性杀菌活性与涉及上述过程的差异调节蛋白数量的增加相关。
获得的结果表明，GO 的时间依赖性杀菌作用归因于其包裹/捕获能力、ROS 产生以及细胞膜的物理破坏。
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