Biophysical Approaches Research Articles

Noncanonical Amino Acid Tools and Their Application to Membrane Protein Studies.

Methods rooted in chemical biology have contributed significantly to studies of integral membrane proteins. One recent key approach has been the application of genetic code expansion (GCE), which enables the site-specific incorporation of noncanonical amino acids (ncAAs) with defined chemical properties into proteins. Efficient GCE is challenging, especially for membrane proteins, which have specialized biogenesis and cell trafficking machinery and tend to be expressed at low levels in cell membranes. Many eukaryotic membrane proteins cannot be expressed functionally in E. coli and are most effectively studied in mammalian cell culture systems. Recent advances have facilitated broader applications of GCE for studies of membrane proteins. First, AARS/tRNA pairs have been engineered to function efficiently in mammalian cells. Second, bioorthogonal chemical reactions, including cell-friendly copper-free "click" chemistry, have enabled linkage of small-molecule probes such as fluorophores to membrane proteins in live cells. Finally, in concert with advances in GCE methodology, the variety of available ncAAs has increased dramatically, thus enabling the investigation of protein structure and dynamics by multidisciplinary biochemical and biophysical approaches. These developments are reviewed in the historical framework of the development of GCE technology with a focus on applications to studies of membrane proteins.

Subtleties in Clathrin heavy chain binding boxes provide selectivity among adaptor proteins of budding yeast

Clathrin forms a triskelion, or three-legged, network that regulates cellular processes by facilitating cargo internalization and trafficking in eukaryotes. Its N-terminal domain is crucial for interacting with adaptor proteins, which link clathrin to the membrane and engage with specific cargo. The N-terminal domain contains up to four adaptor-binding sites, though their role in preferential occupancy by adaptor proteins remains unclear. In this study, we examine the binding hierarchy of adaptors for clathrin, using integrative biophysical and structural approaches, along with in vivo functional experiments. We find that yeast epsin Ent5 has the highest affinity for clathrin, highlighting its key role in cellular trafficking. Epsins Ent1 and Ent2, crucial for endocytosis but thought to have redundant functions, show distinct binding patterns. Ent1 exhibits stronger interactions with clathrin than Ent2, suggesting a functional divergence toward actin binding. These results offer molecular insights into adaptor protein selectivity, suggesting they competitively bind clathrin while also targeting three different clathrin sites.

An Investigation of RNA Methylations with Biophysical Approaches in a Cervical Cancer Cell Model

RNA methylation adds a second layer of genetic information that dictates the post-transcriptional fate of RNAs. Although various methods exist that enable the analysis of RNA methylation in a site-specific or transcriptome-wide manner, whether biophysical approaches can be employed to such analyses is unexplored. In this study, Fourier-transform infrared (FT-IR) and circular dichroism (CD) spectroscopy are employed to examine the methylation status of both synthetic and cellular RNAs. The results show that FT-IR spectroscopy is perfectly capable of quantitatively distinguishing synthetic m6A-methylated RNAs from un-methylated ones. Subsequently, FT-IR spectroscopy is successfully employed to assess the changes in the extent of total RNA methylation upon the knockdown of the m6A writer, METTL3, in HeLa cells. In addition, the same approach is shown to accurately detect reduction in total RNA methylation upon the treatment of HeLa cells with tumor necrosis factor alpha (TNF-α). It is also demonstrated that m1A and m6A methylation induce quite a distinct secondary structure on RNAs, as evident from CD spectra. These results strongly suggest that both FT-IR and CD spectroscopy methods can be exploited to uncover biophysical properties impinged on RNAs by methyl moieties, providing a fast, convenient and cheap alternative to the existing methods.

Identification of new potential inhibitors of pteridine reductase-1 (PTR1) via biophysical and biochemical mechanism-based approaches: Step towards the treatment of Leishmaniasis

Leishmaniasis is a parasitic disease, which spreads from the bite of an infected Phlebotomine fly to human hosts. The disease is characterized by a number of clinical manifestations, such as ulcerative lesions at the site of sandfly bite (cutaneous form), inflammation of mucosal membranes (mucosal leishmaniasis) or the deadly visceral form. This study was aimed to target pteridine reductase-1 (PTR1), a member of short chain dehydrogenases, which accounts for the reduction of conjugated and unconjugated pterins in Leishmania parasite. The ptr1-pET28a+-tev construct was expressed using BL21 (DE3) cells, followed by two tandem purification steps including affinity and gel permeation chromatography. In the next phase, functional studies of PTR1 were performed via screening of an in-house library of 500 compounds. The biochemical-mechanism based assay of PTR1 identified 11 hits that were also found to be non-cytotoxic against human fibroblast cell line (BJ) (except compound 6), and thus further studied via computational technique and saturation transfer difference-nuclear magnetic resonance (STD-NMR) spectroscopy. These high throughput techniques identified six compounds 2, 4, 5, 7, 9, and 11 as active, which were then assessed via in-vitro assay. Among them, compounds 2, 4, and 7 showed substantial leishmanicidal activity, comparable to the standard drug, miltefosine (IC50 value: 31.8 ± 0.2 μM). These results narrowed down the search to 3 compounds as potential leads, with prominent protein-ligand interaction profiles. Hence, the respective compounds can be further assessed for their therapeutic potential against leishmaniasis.

Probing host guest inclusion complex and its applications by biophysical approach subsequently optimized by molecular docking

Herein, we explored the construction of a supramolecular encapsulated complex between Nile blue (NB) and p-sulfonatothiacalix[4]arene (TSC4X). The developed inclusion complex (NB-TSC4X) was established by fluorescence spectroscopy, TGA, FTIR, 1H NMR, and DFT studies. Benesi-Hildebrand calculation showed a linear plot that indicated a 1:1 stoichiometric ratio having a fairly high stability constant of 2720 M−1 in the solution phase. DFT analysis helps us to find out the optimized structure of the inclusion complex. Finally, the binding interaction of the inclusion complex with bovine serum albumin (BSA) was evaluated. In brief, this work uncloses a new strategy to enhance the performance of fluorescent dye.

G-quadruplex formation in RNA aptamers selected for binding to HIV-1 capsid.

HIV-1 capsid protein (CA) is essential for viral replication and interacts with numerous host factors to facilitate successful infection. Thus, CA is an integral target for the study of virus-host dynamics and therapeutic development. The multifaceted functions of CA stem from the ability of CA to assemble into distinct structural components that come together to form the mature capsid core. Each structural component, including monomers, pentamers, and hexamers, presents a variety of solvent-accessible surfaces. However, the structure-function relationships of these components that facilitate replication and virus-host interactions have yet to be fully elucidated. A major challenge is the genetic fragility of CA, which precludes the use of many common methods. To overcome these constraints, we identified CA-targeting aptamers with binding specificity for either the mature CA hexamer lattice alone or both the CA hexamer lattice and soluble CA hexamer. To enable utilization of these aptamers as molecular tools for the study of CA structure-function relationships in cells, understanding the higher-order structures of these aptamers is required. While our initial work on a subset of aptamers included predictive and qualitative biochemical characterizations that provided insight into aptamer secondary structures, these approaches were insufficient for determining more complex non-canonical architectures. Here, we further clarify aptamer structural motifs using focused, quantitative biophysical approaches, primarily through the use of multi-effective spectroscopic methods and thermodynamic analyses. Aptamer L15.20.1 displayed particularly strong, unambiguous indications of stable RNA G-quadruplex (rG4) formation under physiological conditions in a region of the aptamer also previously shown to be necessary for CA-aptamer interactions. Non-canonical structures, such as the rG4, have distinct chemical signatures and interfaces that may support downstream applications without the need for complex modifications or labels that may negatively affect aptamer folding. Thus, aptamer representative L15.20.1, containing a putative rG4 in a region likely required for aptamer binding to CA with probable function under cellular conditions, may be a particularly useful tool for the study of HIV-1 CA.

Interferon gamma-gaillardin nanocomplex formation with improved anti-melanoma effects

This study presents a comprehensive analysis of IFN-γ-Gaillardin nanoparticles (NPs) using a combination of computational, biophysical and cell-based approaches. The molecular docking analysis revealed that both hydrogen and hydrophobic forces are involved in the formation of IFN-γ-Gaillardin complex The interaction between IFN-γ and Gaillardin was further characterized by a pronounced ANS fluorescence spectrum peak and a higher intensity for IFN-γ. The Langmuir, Scatchard, and Hill analyses revealed a higher affinity and lower dissociation constant for IFN-γ NPs compared to IFN-γ alone, suggesting enhanced complex stability. Thermal gravimetric analysis confirmed that the Gaillardin interaction might improve the thermal stability of the NPs. The NPs demonstrated robust stability in various media, highlighting their potential as a delivery system. However, size increase in deionized water suggests the need for formulation optimization. Cell-based assays revealed selective cytotoxicity towards A-375 melanoma cancer cells with minimal impact on non-cancerous HaCaT cells, indicating targeted antitumor effects. Real-time PCR showed gene expression changes consistent with antitumor activity and immune response modulation. The findings suggest that IFN-γ-Gaillardin NPs have potent antitumor properties and the ability to modulate the immune system, warranting further investigation into their therapeutic applications. The development of an IFN-γ-based nanocarrier system for Gaillardin delivery offers a promising approach to melanoma therapy, setting a new direction for NP-based cancer treatment strategies.

Bilateral cellular flows display asymmetry prior to left-right organizer formation in amniote gastrulation.

Bilaterians are defined by a bilaterally symmetrical body plan. Vertebrates exhibit external bilateral symmetry but display left-right (LR) asymmetry in their internal organs. Amniote embryos switch patterning of internal organs from bilateral symmetry to LR-asymmetry, but the initiation of LR symmetry breaking is not well understood. Using chick embryo as a model system due to its easy accessibility and similarity to human development, our biophysical approaches to quantify cellular flows inferred that LR symmetry breaking occurs prior to the formation of Hensen's node, a LR organizer, which serves as a signaling center for LR patterning-gene programs. Our work demonstrates that quantitative biophysical parameters can help unravel the initiation of LR symmetry breaking, suggesting involvement of physical mechanisms in this critical biological patterning process.

Mechanism and regulation of kinesin motors.

Kinesins are a diverse superfamily of microtubule-based motors that perform fundamental roles in intracellular transport, cytoskeletal dynamics and cell division. These motors share a characteristic motor domain that powers unidirectional motility and force generation along microtubules, and they possess unique tail domains that recruit accessory proteins and facilitate oligomerization, regulation and cargo recognition. The location, direction and timing of kinesin-driven processes are tightly regulated by various cofactors, adaptors, microtubule tracks and microtubule-associated proteins. This Review focuses on recent structural and functional studies that reveal how members of the kinesin superfamily use the energy of ATP hydrolysis to transport cargoes, depolymerize microtubules and regulate microtubule dynamics. I also survey how accessory proteins and post-translational modifications regulate the autoinhibition, cargo binding and motility of some of the best-studied kinesins. Despite much progress, the mechanism and regulation of kinesins are still emerging, and unresolved questions can now be tackled using newly developed approaches in biophysics and structural biology.

Novel imaging and biophysical approaches to study tissue hydraulics in mammalian folliculogenesis

AbstractA key developmental stage in mammalian folliculogenesis is the formation of a fluid-filled lumen (antrum) prior to ovulation. While it has long been speculated that the follicular fluid is essential for oocyte maturation and ovulation, little is known about the morphogenesis and the mechanisms driving the antrum formation and ovulation, potentially due to challenges in imaging tissue dynamics in large tissues. Misregulation of such processes leads to anovulation, a hallmark of infertility in ageing and diseases such as the polycystic ovary syndrome (PCOS). In this review, we discuss recent advances in deep tissue imaging techniques, machine learning and theoretical approaches that have been applied to study development and diseases. We propose that an integrative approach combining these techniques is essential for understanding the physics of hydraulics in follicle development and ovarian functions.

Sumoylation of thymine DNA glycosylase impairs productive binding to substrate sites in DNA

The base excision repair enzyme thymine DNA glycosylase (TDG) protects against mutations by removing thymine or uracil from guanine mispairs and functions in active DNA demethylation by excising 5-formylcytosine (fC) and 5-carboxylcytosine (caC). Post-translational modification of TDG by SUMO (small ubiquitin-like modifier) reduces its glycosylase activity but the mechanism remains unclear. We investigated this problem using biochemical and biophysical approaches and a TDG construct comprising residues 82-340 (of 410) that includes the SUMOylation site and the motif for non-covalent SUMO binding. Single turnover kinetics experiments were collected at multiple enzyme concentrations ([E]) and the hyperbolic dependence of activity (kobs) on [E] yielded the maximal glycosylase activity (kmax), the enzyme concentration giving half-maximal activity (K0.5), and the catalytic efficiency (kmax/K0.5). Sumoylation of TDG (or TDG82-340) causes large reductions in catalytic efficiency for G·T, G·U, G·fC, and G·caC DNA substrates, due largely to weakened substrate affinity (increased K0.5). 19F NMR experiments show that sumoylation of TDG82-340 reduces productive binding to G·U mispairs and dramatically impairs binding to G·T mispairs. A mutation in the TDG SUMO-interacting motif (SIM), E310Q, shown previously to perturb noncovalent binding of SUMO to unmodified TDG, rescues the glycosylase activity of sumoylated TDG82-340. Similarly, NMR studies show the mutation restores productive binding of sumoylated TDG82-340 to G·U and G·T pairs. Together, the results indicate that intramolecular SUMO-SIM interactions mediate the adverse effect of sumoylation on TDG activity and suggest a model whereby the disruption of SUMO-SIM interactions enables productive binding of sumoylated TDG to substrate sites in DNA.

Exploring nonlinear dynamics in brain functionality through phase portraits and fuzzy recurrence plots.

Much of the complexity and diversity found in nature is driven by nonlinear phenomena, and this holds true for the brain. Nonlinear dynamics theory has been successfully utilized in explaining brain functions from a biophysics standpoint, and the field of statistical physics continues to make substantial progress in understanding brain connectivity and function. This study delves into complex brain functional connectivity using biophysical nonlinear dynamics approaches. We aim to uncover hidden information in high-dimensional and nonlinear neural signals, with the hope of providing a useful tool for analyzing information transitions in functionally complex networks. By utilizing phase portraits and fuzzy recurrence plots, we investigated the latent information in the functional connectivity of complex brain networks. Our numerical experiments, which include synthetic linear dynamics neural time series and a biophysically realistic neural mass model, showed that phase portraits and fuzzy recurrence plots are highly sensitive to changes in neural dynamics and can also be used to predict functional connectivity based on structural connectivity. Furthermore, the results showed that phase trajectories of neuronal activity encode low-dimensional dynamics, and the geometric properties of the limit-cycle attractor formed by the phase portraits can be used to explain the neurodynamics. Additionally, our results showed that the phase portrait and fuzzy recurrence plots can be used as functional connectivity descriptors, and both metrics were able to capture and explain nonlinear dynamics behavior during specific cognitive tasks. In conclusion, our findings suggest that phase portraits and fuzzy recurrence plots could be highly effective as functional connectivity descriptors, providing valuable insights into nonlinear dynamics in the brain.

Cm-p5, a molluscan-derived antifungal peptide exerts its activity by a membrane surface covering in a non-penetrating mode

Amidst the health crisis caused by the rise of multi-resistant pathogenic microorganisms, Antimicrobial Peptides (AMPs) have emerged as a potential alternative to traditional antibiotics. In this sense, Cm-p5 is an AMP with fungistatic activity against the yeast Candida albicans. Its antimicrobial activity and selectivity have been well characterized; however, the mechanism of action is still unknown. This study used biophysical approaches to gain insight into how this peptide exerts its activity. Stability and fluidity of lipid membrane were explored by liposome leakage and Laurdan generalized polarization (GP) respectively, suggesting that Cm-p5 does not perturb lipid membranes even at very high concentrations (≥100µm.L-1). Likewise, no depolarizing action was observed using 3,3'-propil-2,2'-thyodicarbocianine, a potential membrane fluorescent reporter, with C. albicans cells or the corresponding liposome models. Changes in liposome size were analyzed by Dynamic Light Scattering (DLS) data, indicating that Cm-p5 covers the vesicular surface slightly increasing liposome hydrodynamic size, without liposome rupture. These results were further corroborated with Langmuir monolayer isotherms, where no significant changes in lateral pressure or area per lipid were detected, indicating little or no insertion. Finally, data obtained from molecular dynamics simulations aligned with in vitro observations, whereby Cm-p5 slightly interacted with the fungal membrane model surface without causing significant perturbation. These results suggest Cm-p5 is not a pore-forming anti-fungal peptide and that other mechanisms of action on the membrane as some limitation of fungal nutrition or receptor-dependent transduction for depressing growth development should be explored.

Emergence of synchronized growth oscillations in filamentous fungi.

Many species of soil fungi grow in the form of branched networks that enable long-range communication and mass flow of nutrient. These networks play important roles in the soil ecosystem as a major decomposer of organic materials. While there have been investigations on the branching of the fungal networks, their long-term growth dynamics in space and time is still not very well understood. In this study, we monitor the spatio-temporal growth dynamics of the plant-promoting filamentous fungus Serendipita indica for several days in a controlled environment within a microfluidic chamber. We find that S. indica cells display synchronized growth oscillations with the onset of sporulation and at a period of 3 h. Quantifying this experimental synchronization of oscillatory dynamics, we show that the synchronization can be recapitulated by the nearest neighbour Kuramoto model with a millimetre-scale cell-cell coupling. The microfluidic set-up presented in this work may aid the future characterization of the molecular mechanisms of the cell-cell communication, which could lead to biophysical approaches for controlling fungi growth and reproductive sporulation in soil and plant health management.

Exploring Nonlinear Dynamics In Brain Functionality Through Phase Portraits And Fuzzy Recurrence Plots.

Much of the complexity and diversity found in nature is driven by nonlinear phenomena, and this holds true for the brain. Nonlinear dynamics theory has been successfully utilized in explaining brain functions from a biophysics standpoint, and the field of statistical physics continues to make substantial progress in understanding brain connectivity and function. This study delves into complex brain functional connectivity using biophysical nonlinear dynamics approaches. We aim to uncover hidden information in high-dimensional and nonlinear neural signals, with the hope of providing a useful tool for analyzing information transitions in functionally complex networks. By utilizing phase portraits and fuzzy recurrence plots, we investigated the latent information in the functional connectivity of complex brain networks. Our numerical experiments, which include synthetic linear dynamics neural time series and a biophysically realistic neural mass model, showed that phase portraits and fuzzy recurrence plots are highly sensitive to changes in neural dynamics and can also be used to predict functional connectivity based on structural connectivity. Furthermore, the results showed that phase trajectories of neuronal activity encode low-dimensional dynamics, and the geometric properties of the limit-cycle attractor formed by the phase portraits can be used to explain the neurodynamics. Additionally, our results showed that the phase portrait and fuzzy recurrence plots can be used as functional connectivity descriptors, and both metrics were able to capture and explain nonlinear dynamics behavior during specific cognitive tasks. In conclusion, our findings suggest that phase portraits and fuzzy recurrence plots could be highly effective as functional connectivity descriptors, providing valuable insights into nonlinear dynamics in the brain.

Exploring the effects of 4-chloro-o-phenylenediamine on human fibrinogen: A comprehensive investigation via biochemical, biophysical and computational approaches

Fibrinogen (Fg), an essential plasma glycoprotein involved in the coagulation cascade, undergoes structural alterations upon exposure to various chemicals, impacting its functionality and contributing to pathological conditions. This research article explored the effects of 4-Chloro-o-phenylenediamine (4-Cl-o-PD), a common hair dye component (IUPAC = 1-Chloro-3,4-diaminobenzene), on human fibrinogen through comprehensive computational, biophysical, and biochemical approaches. The formation of a stable ligand-protein complex is confirmed through molecular docking and molecular dynamics simulations, revealing possible interaction having a favorable −4.8 kcal/mol binding energy. Biophysical results, including UV–vis and fluorescence spectroscopies, corroborated with the computational findings, whereas Fourier transform infrared spectroscopy (FT-IR) and circular dichroism spectroscopy (CD) provide insights into the alterations of secondary structures upon interaction with 4-Cl-o-PD. Anilinonaphthalene-sulfonic acid (ANS) fluorescence showed a partially unfolded protein, with enhanced α to β-sheet transition as evidenced by thioflavin T (ThT) spectroscopy and microscopy. Moreover, biochemical assays confirmed the formation of carbonyl compounds that may be responsible for the oxidation of methionine residues in fibrinogen. Electrophoresis and electron microscopy confirmed the formation of aggregates. Our findings elucidate the interaction pattern of 4-Cl-o-PD with Fg, leading to structural perturbation, which may have potential implications for fibrinogen misfolding or its aggregation. Protein aggregation or its misfolded products affect peripheral tissues and the central nervous system. Many chronic progressive diseases, like type II diabetes mellitus, Alzheimer's disease, Parkison's disease, and Creutzfeldt-Jakob disease are associated with intrinsically aberrant disordered proteins. Understanding these interactions may offer new perspectives on the safety and biocompatibility of dye compounds, which may contribute to developing improved strategies for acquired amyloidogenesis.

Static magnetic field effects on the secondary structure and elasticity of collagen molecules; a possible biophysical approach to treat keratoconus

Type I collagen is among the major extracellular proteins that play a significant role in the maintenance of the cornea's structural integrity and is essential in cell adhesion, differentiation, growth, and integrity. Here, we investigated the effect of 300 mT Static Magnetic Field (300 mT SMF) on the structure and molecular properties of acid-solubilized collagens (ASC) isolated from the rat tail tendon. The SMF effects at molecular and atomic levels were investigated by various biophysical approaches like Circular Dichroism Spectropolarimetery (CD), Fourier Transform Infrared Spectroscopy (FTIR), Zetasizer light Scattering, and Rheological assay. Exposure of isolated type I collagen to 300 mT SMF retained its triple helix. The elasticity of collagen molecules and the keratoconus (KCN) cornea treated with SMF decreased significantly after 5 min and slightly after 10, 15, and 20 min of treatments. The exposure to 300 mT SMF shifted the Amid I bond random coil to antiparallel wave number from 1647 to 1631 cm−1. The pH of the 300 mT SMF treated collagen solution increased by about 25 %. The treatment of the KCN corneas with 300 mT SMF decreased their elasticity significantly. The promising results of the effects of 300 mT SMF on the collagen molecules and KCN cornea propose a novel biophysical approach capable of manipulating the collagen's elasticity, surface charges, electrostatic interactions, cross binding, network formation and fine structure. Therefore, SMF treatment may be considered as a novel non-invasive, direct, non-chemical and fast therapeutic and manipulative means to treat KCN cornea where the deviated physico-chemical status of collagen molecules cause deformation.

Label-free live characterization of mesenchymal stem cell spheroids by biophysical properties measurement

Three-dimensional (3D) cell culture has become a consolidated method in the stem cell field, where mesenchymal stromal stem cells (MSCs) can be used to generate in vitro spheroid aggregates called MSC-Spheroids (MSph). MSph is a floating cluster of stem cells similar to those in literature are known as bone marrow-derived “mesenspheres”. Even though MSC properties are shared by MSph, depending on the cell type and their tissue source, the morphology and degree of compaction of the MSph can be variable, creating limitations in such a cell model. Thus, during culture, a variation in stem cell functionality and viability, in addition to the suitability for comparing MSph in some experimental protocols, can be affected by spheroid biophysical intrinsic properties like mass density. To investigate this limitation and provide a new method for researchers, MSph of seven different tissue sources were compared by combining mass density, weight, and size evaluations with viability assays for ATP measurement. MSph cultured in traditional static conditions showed decreased in viability over the days of culture, while mass density exhibited different trends among cell types. Additionally, treatment of MSph with a non-toxic concentration of a natural compound cell regulator, such as plumbagin, altered the mass density of a selected cell type, thereby confirming the efficacy of the biophysical approach in monitoring MSph variability post-treatment. The results encourage using MSph in the early days of culture after their formation to ensure viability and likely retention of the stem cell phenotype.

How bacteria actively use passive physics to make biofilms

Modern molecular microbiology elucidates the organizational principles of bacterial biofilms via detailed examination of the interplay between signaling and gene regulation. A complementary biophysical approach studies the mesoscopic dependencies at the cellular and multicellular levels with a distinct focus on intercellular forces and mechanical properties of whole biofilms. Here, motivated by recent advances in biofilm research and in other, seemingly unrelated fields of biology and physics, we propose a perspective that links the biofilm, a dynamic multicellular organism, with the physical processes occurring in the extracellular milieu. Using Bacillus subtilis as an illustrative model organism, we specifically demonstrate how such a rationale explains biofilm architecture, differentiation, communication, and stress responses such as desiccation tolerance, metabolism, and physiology across multiple scales—from matrix proteins and polysaccharides to macroscopic wrinkles and water-filled channels.

Insight into the Binding Interaction Mechanism of the Ligand M1069 with Human Serum Albumin and A2A Adenosine Receptor—A Biophysical Approach

The study focuses on M1069, a promising drug for solid tumors functioning as an A2A adenosine receptor antagonist. Herein, we investigate the binding mechanism of the drug molecule M1069 by employing computational techniques to draw comparisons with Human serum albumin (HSA), shedding light on the intricate interactions between M1069 and its target as well. The molecular docking results suggest that the drug molecule binds very well with the HSA protein when compared to A2AAR with the docking score of −7.02 kcal/mol. To substantiate the molecular interactions between the drug and the protein structure, comprehensive molecular dynamic simulations were performed for a period of 100 ns. This approach facilitated an in-depth analysis of the dynamic relationships and behaviors between the drug and the protein, offering valuable insights into their intermolecular dynamics and stability. Subsequently, DFT analysis is conducted to assess the inhibitory efficacy of the drug, employing the B3LYP technique. Additionally, an investigation into the pharmacological ADMET parameters of the drug molecule resulted in gastrointestinal absorption. These outcomes contribute to a better understanding of the interaction dynamics between the drug and the proteins, providing valuable insights that can enhance the drug discovery process.