Modares Journal of Biotechnology

Abstract Ferroptosis is a newly identified form of cell death associated with lipid peroxidation. This process is dependent on iron and polyunsaturated fatty acids (PUFAs). Despite the importance of ferroptosis, the molecular details of this process, particularly its impact on cellular membrane properties, remain unknown. In this study, structural and permeability changes in the plasma membrane resulting from lipid peroxidation during ferroptosis were investigated using molecular dynamics simulations. Initially, a model of the human red blood cell membrane was constructed based on experimental data. To simulate ferroptosis, the PUFA lipid chains in the red blood cell membrane were replaced with their hydroperoxide derivatives. Both systems (normal and ferroptotic membranes) were examined in All-Atom molecular dynamics simulations for 300 nanoseconds (with three replicates). The results showed that in the ferroptotic membrane, the thickness decreased, and the surface area increased. Additionally, the hydroperoxide groups in the fatty acid chains moved toward the polar head groups of the phospholipids. Besides these structural changes, the function of the membrane, which typically acts as an impermeable barrier to polar molecules such as water, was disrupted due to lipid peroxidation, while the overall membrane integrity remained intact. In summary, lipid peroxidation in ferroptosis induces significant changes in membrane structure and function, which could be utilized in the development of new treatments for severe diseases such as cancer and neurodegenerative disorders.

Abstract Today, the engineering of bone tissue has created special solutions to restore bone tissue by combining biological materials with a scaffold to provide cells suitable for bone formation and growth factors. In this research, a fusion peptide was designed with bioinformatics methods that can bind to the growth factors involved in bone tissue repair and lead to the trapping of these factors in the lesion site. In this study, heparin-binding domain was placed in the designed peptide and this peptide was complexed with growth factor in monomer and dimer forms with the help of docking. The structures of the complex were selected based on the lowest scores obtained, which included -912.5 and -1117, respectively. According to the results of docking and molecular dynamics simulation, this fusion peptide was able to bind to the growth factor of bone morphogenetic proteins. Based on the results of the simulation, unlike the peptide in the monomer state, the changes in the RMSD diagram of the peptide complex in the dimer state became stable after 10 nanoseconds from the simulation time and remained stable until the end of the simulation. These results show that the resulting complex in the dimer state has a better pattern of stability compared to the monomer state according to the investigation of the RMSD factor.

Abstract Abstract

The increase in antibacterial resistance due to the high consumption of antibiotics is a looming crisis for humanity. Natural antibacterial compounds, such as cationic antibacterial peptides, hold a significant position, as the likelihood of developing resistance against them is low. CAP37, or AZU1, is a protein derived from the granules of human neutrophils and functions as a natural antibiotic. The residues 20-44 of CAP37 exhibit antibacterial activity. In this study, two mutations were designed in the native CAP37 peptide to enhance its antibacterial activity. Molecular dynamics simulations of the peptides in water were conducted for 50 ns, along with umbrella sampling MD simulations to obtain reliable potential of mean force (PMF) profiles for each peptide's passage through lipid A, a component of the bilayer membrane. The results indicated that the peptide containing the SWRW sequence exhibited lower aggregation properties in water and a greater tendency to traverse the lipid A bilayer membrane compared to the native (SQRS) or WWRS sequences. Therefore, it can be concluded that CAP37 with the SWRW sequence has enhanced antimicrobial activity. To validate the theoretical findings, both the natural and SWRW mutant peptides were synthesized. The minimum inhibitory concentration (MIC) test conducted on various Gram-negative and Gram-positive bacteria, including Escherichia coli, Klebsiella pneumoniae, Staphylococcus aureus, and Bacillus subtilis, demonstrated that the mutated peptide exhibited antibacterial activity, thereby confirming the theoretical results.

Abstract Viral infections represent pathological conditions arising from the intrusion of viruses into host cells and their replication. The onset of infection is intricately tied to the interplay between viral and host cell proteins. Thus, elucidating these protein-protein interactions assumes a pivotal role in the encompassing prevention, treatment, and control of viral infections. Given traditional laboratory experimentation's prohibitively high costs and time-intensive nature, researchers have increasingly turned to computational approaches for predicting human-virus protein-protein interactions. Despite the performance of these computational approaches, a challenge persists in the need for an effective protein representation that adequately captures their structural intricacies.
In this paper, we present PBS, a novel model for the prediction of protein-protein interactions between viruses and humans. PBS leverages the transformers to effectively represent proteins. The model unified the latent space for human and virus proteins through the implementation of heterogeneous siamese neural networks.The model achieves an accuracy score of 81.41%, an area under the ROC curve score of 87.35%, an area under the precision-recall curve score of 87.78%, an F1 score of 81.58%, and a precision score of 80.84%. These metrics collectively underscore the satisfactory performance of the PBS model.
Furthermore, we assess the model's predictive capabilities in discerning interactions between proteins associated with the H1N1 influenza virus and human proteins.

Abstract Introduction: Macrophages are considered a particularly challenging cell type to transfect. Given their importance as therapeutic targets, developing a successful transfection method for these cells is highly desirable.
Materials and Methods: The efficiency of lentiviral transfection was compared to three commercially transfection reagents (Xfect™ Transfection Reagent, FuGENE® HD, and Lipofectamine ^TM 3000) in RAW264.7 macrophage cells. Following optimization and production of lentiviral particles in 293T cells, RAW264.7 cells were infected with varying MOI. Transduction efficiency, cell viability, and metabolic activity were measured and compared to the transfection efficiency of the chemical methods.
Results: None of the three chemical transfection reagents successfully transfected RAW264.7 cells. In contrast, the lentiviral method achieved transduction even at the lowest concentration of viral stock, with a green fluorescent signal observable under a fluorescence microscope. Increasing the viral stock concentration and using higher MOIs (up to 30) significantly (p≤0.0001) increased transduction efficiency.
Discussion: Despite requiring more time and effort than chemical methods, lentiviral transduction exhibited superior efficiency in transfecting hard-to-transfect cells and further improvements were achieved through some modifications such as virus concentration, the use of polybrene, no viral freezing and O/N incubation with concentrated viral particles. Since other parameters, especially the use of retronectin and spinoculation, are effective on the efficiency of the virus infection process, it is suggested that they be considered in future studies and given the encouraging data of this research, this completed method can also be applied to other difficult-to-transfect cells, such as different types of stem cells or primary cells.

Abstract Cancer is one of the causes of death in human societies, and the main reason for failure in its treatment is late diagnosis and involvement of other body organs. Biomarkers measurement of body fluids can be one of the most important methods of cancer screening and diagnosis in the early stages. Circulating microRNAs have been proposed as new biomarkers for cancer diagnosis and prognosis. The use of these molecules, in addition to early and timely diagnosis before metastasis, will reduce the damage to the patient due to the possibility of non-invasive access. Therefore, developing a method to identify, reveal and quantify it is a necessity. In this study, a biosensor was designed to isolate and identify circulating blood microRNAs in a hydrogel platform. In our work, microRNA was isolated using a single-stranded DNA receptor probe and fixed in a platform containing hydrogels. By attaching the microRNA to the probe, the second probe, which complements the biotinylated DNA at the top of the microRNA, forms a sandwich structure. Finally, microRNA trapped between the two probes was detected by FITC-bound streptavidin in the hydrogel platform. In this research, different concentrations of microRNA-9 from 1000 picomolar to 1 femtomol were used to estimate the limit of detection (LOD) of the designed biosensor, and 50 femtomol was measured as the detection limit.

Abstract

Abstract In this study, the electrostatic interactions between the third and fourth antiparallel helices of Mnemiopsin 2 have studied. Due to the distribution of the positive and negatively charged residues at the C- and N-terminals of helices, we treated them as two antiparallel nanowires with electric dipole character. By mutating Phe87 to Arg, we aimed to enhance the electric dipole of the fourth helix. The MODELLER program was employed to generate the tertiary structure of the wild-type and mutant proteins. The expression system was prepared using the site-directed mutagenesis procedure. Activity measurements revealed that while the wild-type protein exhibited faster reaction initiation and a higher decay rate. However, the mutant displayed a significantly higher photon yield, approximately double that of the wild-type. Intrinsic fluorescence measurements indicated that the microenvironment around the chromophores in the mutant photoprotein had altered, leading to increased distance of chromophores from internal quenchers. However, the overall structure of the mutant protein became more compact, as confirmed by the ANS-based fluorescence. Heat-induced denaturation experiments showed that while the melting temperature (T_m) remained unaffected, the enthalpy change of denaturation increased significantly in the mutant, suggesting enhanced cooperativity in the intramolecular stabilizing interactions. Finally, it was concluded that increasing the cooperativity between the stabilizing interactions in the mutant protein, stabilize the native structure, leading to a higher population of functional complexes and, consequently, a higher photon yield.