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    Cross-reacting material 197 (CRM197) affects actin cytoskeleton of endothelial cells
    (General Physiol And Biophysics, 2017) Edis, Bilge Ozerman; Varol, Basak; Haciosmanoglu, Ebru; Unlu, Ayhan; Bektas, Muhammet
    CRM197, cross-reacting material 197, is a mutant of diphtheria toxin (DTx). CRM197 is used in pharmacology as a carrier protein. It has been recently shown that CRM197 causes breakdown in actin filaments. In order to show intracellular localization of CRM197 and visualize cell structure via actin cytoskeleton, endothelial cells were cultured and subjected to CRM197 in vitro. To address the interaction between CRM197 and actin both experimental and theoretical studies were carried out. Colocalization of CRM197 with actin filaments was determined by immunofluorescence microscopy. Following 24-hour incubation, the loss of cell-cell contact between cells was prominent. CRM197 was shown to bind to G-actin by gel filtration chromatography, and this binding was confirmed by Western blot analysis of eluted samples obtained following chromatography. Based on crystal structure, docked model of CRM197-actin complex was generated. Molecular dynamics simulation revealed that Lys42, Cys218, Cys233 of CRM197 interacts with Gly197, Arg62 and Ser60 of G-actin, respectively. CRM197 binding to G-actin, colocalization of CRM197 with actin filament, and actin cytoskeleton rearrangement resulting in the loss of cell-cell contact show that actin comes into sight as target molecule for CRM197.
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    Noncovalent Pyrene-Polyethylene Glycol Coatings of Carbon Nanotubes Achieve in Vitro Biocompatibility
    (AMER CHEMICAL SOC, 2018) Meran, Mehdi; Akkus, Pelin Deniz; Kurkcuoglu, Ozge; Baysak, Elif; Hizal, Gurkan; Haciosmanoglu, Ebru; Unlu, Ayhan; Karatepe, Nilgun; Guner, F. Seniha
    Single-walled carbon nanotubes (SWNTs) have become increasingly exploited in biological applications, such as imaging and drug delivery. The application of SWNTs in biological settings requires the surface chemistry to remain through the low solubility in aqueous media. In this research, a facile approach for the preparation of a polyethylene glycol (PEG)-coated SWNT-based nanocarrier was reported. We focused on the effect of PEG chain length and SWNT size on the cytotoxicity of PEG coated SWNTs as a superior drug delivery nanovector. First, all-atom molecular dynamics (MD) simulations were employed to explore the stability and behavior of SWNT/pyrene-PEG (SWNT/Pyr-PEG) structures at a molecular level that is not attainable with experiments. The MD studies revealed that (i) a pi-pi stacking interactions between the pyrene bearing PEG molecules and SWNTs are maintained in bulky situations, regardless of PEG molecular weight or SWNT size; (ii) pyrene molecules diffuse over the SWNT surface without detaching; and (iii) both short and long dynamic Pyr-PEG chains have the capability of effectively coating the SWNT surface. In light of the simulations, noncovalent (pi-pi stacking) assemblies of SWNT/Pyr-PEG with different molecular weights of PEG (M-W = 2000, 5000, and 12000) were successfully fabricated and characterized. For longer PEG chains, more effective coating of SWNTs was obtained, resulting in more biocompatible SWNT/Pyr-PEG nanomaterials. The number of SWNTs coated by Pyr-PEG was highly dependent on the length of pyrene bearing PEG polymers. Moreover, the short SWNTs showed a higher amount of PEG coating with respect to the long SWNTs. Cell viability results demonstrated a dose-dependent cytotoxicity of coated SWNTs. Short SWNTs coated with longer PEG chains have low cytotoxicity to be used in in vivo studies.