Biophysical, bioinformatical, cellular, and molecular investigations on the effects of graphene oxide nanosheets on the hemoglobin structure and lymphocyte cell cytotoxicity

Abstract

BackgroundImplementations of nanoparticles have been receiving great interest in medicine and technology due to their unique characteristics. However, their toxic impacts on the biological system are not well explored.AimThis study aims to investigate the influence of fabricated nano graphene oxide (NGO) sheets on the secondary and quaternary structural alterations of human hemoglobin (Hb) and cytotoxicity against lymphocyte cells.Materials and methodsDifferent spectroscopic methods, such as extrinsic and synchronous fluorescence spectroscopy and far circular dichroism (CD) spectroscopy, molecular docking investigation, cellular assays (trypan blue exclusion, cellular uptake, ROS, cell cycle, and apoptosis), and molecular assay (fold changes in anti/proapoptotic genes [B-cell lymphoma-2 {BCL2}/BAX] expression levels) were used in this study.ResultsTransmission electron microscopy, X-ray diffraction, Fourier transform infrared spectroscopy, and zeta potential investigations revealed the nano-sized nature of NGOs with good colloidal stability. Extrinsic fluorescence spectroscopy by using 8-anilinonaphthalene-1 -sulfonic acid and synchronous fluorescence spectroscopy showed that NGOs can unfold the quaternary structure of Hb in the vicinity of Tyr residues. The CD investigation demonstrated that the α-helicity of Hb experienced substantial alteration upon interaction with increasing concentrations of NGOs. The molecular docking study showed that NGOs interacted with polar residues of Hb. Cellular and molecular assays revealed that NGOs lead to ROS formation, cell cycle arrest, and apoptosis through the BAX and BCL2 pathway.ConclusionThese data reveal that NGOs can induce some protein structural changes and stimulate cytotoxicity against normal cell targets. Therefore, their applications in healthy systems should be limited.

Nano-Graphene Oxide-Promoted Epithelial–Mesenchymal Transition of Human Retinal Pigment Epithelial Cells through Regulation of Phospholipase D Signaling

“…Nano-GO has been reported to upregulate oxidative stress in macrophages, lymphocytes, and embryonic fibroblast cells [39,40]. Normal ROS production is essential for maintaining normal physiological functions in human RPE cells.…”

<p>Cerium oxide NPs mitigate the amyloid formation of α-synuclein and associated cytotoxicity</p>

“…The expression amount of (BCL-2) and BAX genes was estimated by real-time PCR analysis based on our previous paper 28. TRIzol reagent (Thermo Fisher Scientific, Waltham, MA, USA) and RevertAid first-strand cDNA synthesis kit (Takara, Japan) were used to extract total RNA and synthesis of cDNA, respectively according to the manufacturer’s instructions.…”

The nanosafety assessment of ENMs under a dermal exposure scenario: from key molecular events to in silico modeling tools

“…For example, graphene oxide (NGO) nano-sheets were docked with human hemoglobin and the results indicate that NGOs can denature the quaternary and secondary structures of proteins in the vicinity of Tyr residues to simulate cytotoxicity against normal cell targets. 190 In another study, the docking predictions of the binding sites, binding affinities, and interaction modes of SiO 2 NPs and TiO 2 NPs confirmed in vitro that the adsorption of NPs causes disturbance in the secondary and tertiary structures of antioxidant enzymes (catalase and superoxide dismutase), resulting in loss of enzyme activity, which provides a new perspective to explain ENM-driven oxidative stress. 195 Likewise, an investigation of the interaction between ZnO NPs and bovine serum albumin (BSA) and human insulin (HI) by molecular docking simulations reveals that the nanostructures of the NPs and the nature of the selected protein are key factors affecting the ability of ZnO NPs in treating amyloid diseases.…”

“…189 As Fig. 6 shows, a typical docking simulation involves three steps: i) preparation (optimization) of the crystal structural information for ENM ligands (obtainable from online resources) and receptor molecules that can bind to ENMs (structures downloaded from the Protein Data Bank or homogenized using SwissModel, Modeler and Prime software); ii) calculation of the binding energy to predict the binding site and binding mode by AutoDock software based on the AMBER force field; iii) visualization of the nano- Table 4 lists the previous molecular docking studies of ENMs focusing on their interactions with various biological macromolecules, e.g., proteins [190][191][192][193] and nucleic acids, 194 after internalized into the human body. Recently, molecular docking simulation has been examined as a tool to obtain the preferred binding modes of nanoparticles and proteins, and the obtained data can reflect the detected in vitro activities or toxicity.…”

“…Table 4 lists the previous molecular docking studies of ENMs focusing on their interactions with various biological macromolecules, e.g. , proteins 190–193 and nucleic acids, 194 after internalized into the human body. Recently, molecular docking simulation has been examined as a tool to obtain the preferred binding modes of nanoparticles and proteins, and the obtained data can reflect the detected in vitro activities or toxicity.…”