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A method for removing time- and radially invariant noise from sedimentation velocity and sedimentation equilibrium experiments performed in an analytical ultracentrifuge is presented. The method averages repeat radial incident light measurements as a function of the photomultiplier response at different wavelengths to remove the majority of the time-invariant noise contributions from intensity data measurements. The results of this method are compared to traditional absorbance data generated with a buffer reference and the Beckman Optima AUC data acquisition program, and with the standard UltraScan refinement workflow. The method avoids the amplification of stochastic noise inherent in the absorbance scan subtraction traditionally employed in sedimentation velocity and equilibrium data. In addition, the collection of intensity data frees up the reference channel for additional samples, doubling the capacity of the instrument. In comparison to absorbance data, the residual mean square deviation of a fitted sedimentation velocity experiment without additional noise correction by UltraScan was improved by a factor of 4.5 when using the new method. This improvement benefits sedimentation equilibrium experiments as well as analytical buoyant density equilibrium experiments where routine time-invariant noise correction calculations cannot be performed., (© 2023. European Biophysical Societies' Association.)

Multi-wavelength analytical ultracentrifugation (MW-AUC) is a recently developed technique that has proven to be a promising tool to investigate mixtures of molecules containing multiple chromophores. It provides an orthogonal separation approach by distinguishing molecules based on their spectral and hydrodynamic properties. Existing software implementations do not permit the user to assess the integrity of the spectral decomposition. To address this shortcoming, we developed a new spectral decomposition residual visualization module, which monitors the accuracy of the spectral decomposition. This module assists the user by providing visual and statistical feedback from the decomposition. The software has been integrated into the UltraScan software suite and an example of a mixture containing thyroglobulin and DNA is presented for illustration purposes., (© 2023. European Biophysical Societies' Association.)

Biomechanical and morphological analysis of the cells is a novel approach for monitoring the environmental features, drugs, and toxic compounds' effects on cells. Graphene oxide (GO) has a broad range of medical applications such as tissue engineering and drug delivery. However, the effects of GO nanosheets on biological systems have not been completely understood. In this study, we focused on the biophysical characteristics of cells and their changes resulting from the effect of GO nanosheets. The biophysical properties of the cell population were characterized as follows: cell stiffness was calculated by atomic force microscopy, cell motility and invasive properties were characterized in the microfluidic chip in which the cells are able to visualize cell migration at a single-cell level. Intracellular actin was stained to establish a quantitative picture of the intracellular cytoskeleton. In addition, to understand the molecular interaction of GO nanosheets and actin filaments, coarse-grained (CG) molecular dynamics (MD) simulations were carried out. Our results showed that GO nanosheets can reduce cell stiffness in MCF7 cells and MDA-MB-231 cell lines and highly inhibited cell migration (39.2%) in MCF-7 and (38.6%) in MDA-MB-231 cell lines through the GO nanosheets-mediated disruption of the intracellular cytoskeleton. In the presence of GO nanosheets, the cell migration of both cell lines, as well as the cell stiffness, significantly decreased. Moreover, after GO nanosheets treatment, the cell actin network dramatically changed. The experimental and theoretical approaches established a quantitative picture of changes in these networks. Our results showed the reduction of the order parameter in actin filaments was 23% in the MCF7 cell line and 20.4% in the MDA-MB-231 cell line. The theoretical studies also showed that the GO nanosheet-actin filaments have stable interaction during MD simulation. Moreover, the 2D free energy plot indicated the GO nanosheet can induce conformational changes in actin filaments. Our findings showed that the GO nanosheets can increase the distance of actin-actin subunits from 3.22 to 3.5 nm and in addition disrupt native contacts between two subunits which lead to separate actin subunits from each other in actin filaments. In this study, the biomechanical characteristics were used to explain the effect of GO nanosheets on cells which presents a novel view of how GO nanosheets can affect the biological properties of cells without cell death. These findings have the potential to be applied in different biomedical applications., (© 2021. The Author(s).)

We aimed to explore and compare new insights on the pharmacological potential of Oliveria decumbence essential oil (OEO) and its main components highlighting their antioxidant activity in-vitro, in-vivo, and in-silico and also cytotoxic effects of OEO against A549 lung cancer cells. At first, based on GC-MS analysis, thymol, carvacrol, p-cymene, and γ-terpinene were introduced as basic ingredients of OEO and their in-vitro antioxidant capacity was considered by standard methods. Collectively, OEO exhibited strong antioxidant properties even more than its components. In LPS-stimulated macrophages treated with OEO, the reduction of ROS (Reactive-oxygen-species) and NO (nitric-oxide) and down-regulation of iNOS (inducible nitric-oxide-synthase) and NOX (NADPH-oxidase) mRNA expression was observed and compared with that of OEO components. According to the results, OEO, thymol, and carvacrol exhibited the highest radical scavenging potency compared to p-cymene, and γ-terpinene. In-silico Molecular-Docking and Molecular-Dynamics simulation indicated that thymol and carvacrol but no p-cymene and γ-terpinene may establish coordinative bonds in iNOS active site and thereby inhibit iNOS. However, they did not show any evidence for NOX inhibition. In the following, MTT assay showed that OEO induces cytotoxicity in A549 cancer cells despite having a limited effect on L929 normal cells. Apoptotic death and its dependence on caspase-3 activity and Bax/Bcl2 ratio in OEO-treated cells were established by fluorescence microscopy, flow cytometry, colorimetric assay, and western blot analysis. Additionally, flow cytometry studies demonstrated increased levels of ROS in OEO-treated cells. Therefore, OEO, despite showing antioxidant properties, induces apoptosis in cancer cells by increasing ROS levels. Collectively, our results provided new insight into the usage of OEO and main components, thymol, and carvacrol, into the development of novel antioxidant and anti-cancer agents., (© 2021. The Author(s).)

Lamellar and hexagonal lipid structures are of particular importance in the biological processes such as membrane fusion and budding. Atomistic simulations of formation of these phases and transitions between them are computationally prohibitive, hence development of coarse-grained models is an important part of the methodological development in this area. Here we apply systematic bottom-up coarse-graining to model different phase structures formed by 1,2-dioleoylphosphatidylethanolamine (DOPE) lipid molecules. We started from atomistic simulations of DOPE lipids in water carried out at two different water/lipid molar ratio corresponding to the lamellar Lα and inverted hexagonal HII structures at low and high lipid concentrations respectively. The atomistic trajectories were mapped to coarse-grained trajectories, in which each lipid was represented by 14 coarse-grained sites. Then the inverse Monte Carlo method was used to compute the effective coarse-grained potentials which for the coarse-grain model reproduce the same structural properties as the atomistic simulations. The potentials derived from the low concentration atomistic simulation were only able to form a bilayer structure, while both Lα and HII lipid phases were formed in simulations with potentials obtained at high concentration. The typical atomistic configurations of lipids at high concentration combine fragments of both lamellar and non-lamellar structures, that is reflected in the extracted coarse-grained potentials which become transferable and can form a wide range of structures including the inverted hexagonal, bilayer, tubule, vesicle and micellar structures., Competing Interests: The authors have declared that no competing interests exist.