Biophysical reviews最新文献

Enzymes are highly efficient natural catalysts with exceptional selectivity, often surpassing synthetic alternatives in accelerating chemical reactions. Their water solubility enables their application in various technological processes aligned with modern green chemistry trends. However, enhancing enzyme performance is often necessary, primarily by improving reusability and extending their operational lifespan. A promising strategy to achieve these goals is enzyme immobilization, particularly on cellulose. Cellulose, the most abundant biopolymer, possesses valuable properties such as cost-effectiveness and low toxicity, making it an ideal candidate for applications in food and biomedical technologies. Additionally, cellulose-based immobilization can contribute to the development of enzyme formulations with improved catalytic efficiency and stability. This review summarizes over two hundred studies conducted in the past decade on enzyme immobilization on cellulose and its derivatives, emphasizing its effects on enzymatic activity and performance.

The transition of immunotherapy administration from intravenous infusion to subcutaneous (SC) administration of monoclonal antibody formulations for oncology patients has garnered significant interest. SC administration offers multiple benefits, including potential for at-home administration, enhanced patient compliance, reduced hospital congestion, lowered health care costs, and improved sustainability by reducing drug wastage and minimizing environmental impact. However, for many biologics, the shift to SC administration requires the development of high-concentration monoclonal antibody products (HCmAP) due to the need for large dose volumes. Here we explore the impact of the COVID-19 pandemic on immunotherapy administration and the imperative of adopting SC administration. We discuss challenges encountered throughout the manufacturing, shipping, storage, and delivery of HCmAP. A central hurdle identified involves the biophysical instability and the large increase in viscosity of these biologics due to increased antibody concentration. Further complications can arise from "non-ideality" effects through molecular crowding or co-exclusion effects (macromolecules blocking the free movement in solution of other macromolecules) and elevated macromolecular interactions. For reducing the viscosity for a given concentration of antibody, the main excipients reported are salts and amino acids, with Arg-HCl demonstrating particularly improved formulation viscosity in an HCmAP. However, excipients with viscosity-lowering effects can also impact protein stability. The journey to discover suitable excipient strategies remains ongoing, combined with emerging approaches such as molecular engineering and computational techniques, with the ultimate aim of facilitating the successful integration of SC administration for economic savings, environmental sustainability, and social equity.

This review describes various molecular biophysics methods (crystallography, mass spectrometry, NMR spectroscopy, electron cryo-microscopy (cryoEM), free electron laser and X-ray photon correlation spectroscopy) which can be used to investigate the molecular structure of proteins under diverse conditions (visually summarised in the thumbnail image for the journal contents pages). We focus particularly on those which permit for the incubation and/or experimental sample analysis at mammalian body temperature (37 °C) or at physiological conditions for yet higher temperatures such as thermophiles. Crystallography, a leading method in structure elucidation, in recent decades has been dominated by structures analysed at cryogenic temperatures to ensure best resolution and crystal stability under X-ray irradiation. However, it raises the question-is the atomic structure elucidated by cryo-crystal structures truly representative of processes occurring at body temperature? This is surely an important requirement for protein-ligand binding investigations for drug discovery as protein binding may vary with temperature and indicate key aspects that could be overlooked. A review of wwPDB submissions versus sample temperature analysis clearly indicates a marked lack of atomic data obtained at 37 °C. This is not to say that 100 K cryo-crystallography ought to be replaced, in such structure-based drug discovery which is highly efficiently implemented at many macromolecular crystallographic beamlines worldwide, but favoured ligand binding events in particular for lead compounds from those surveys may provide additional valued data when studied at 37 °C.

Graphical abstract: Thumbnail contents image: Molecular biophysics methods suitable for the analysis of macromolecules at body temperatures or higher.

Supplementary information: The online version contains supplementary material available at 10.1007/s12551-025-01328-4.

The investigation of the interactions of small lipophilic molecules (e.g., drugs) with lipid membranes represents an active field of contemporary biophysical research. The determination of their membrane insertion, their distribution within the lipid bilayer, and their effect on lipid membranes themselves hold significant pharmacological importance since the plasma membrane often represents the first contact site for the interaction of a drug with the cell. In this work, we review recent applications of solid-state NMR spectroscopy that have been conducted to study the interaction of lipid membranes with a large variety of small drug-like molecules (e.g., local anesthetics, statins, NSAIDs, kinase inhibitors). We aim to briefly highlight previous research while outlining the promising prospects using experimental and computational methods, including ²H, ³¹P, ¹H magic-angle spinning (MAS) NOESY NMR, and molecular dynamics (MD) simulations, which provide highly useful tools for a comprehensive understanding of these interactions.

We review current trends of courses and young scientists' schools on bioinformatics discussed at the Congress of Russian Biophysicists and the recent Russian Autumn School in Biophysics held in Kazan, Russia, 2024, highlighting general problems in educational courses organization. We discuss the history of bioinformatics educational courses and meetings and current trends in bioinformatics education at regional and international levels. The distant education forms in bioinformatics courses and the approaches development active learning. Here, we discuss the educational courses on bioinformatics, the integration of online tools, gamification conception, and AI tools for self-education.

The main motivation of this work was to address the challenge of single-molecule functional study of membrane proteins under stable and independently controlled electrical and chemical membrane potentials. Although transmembrane potential is often essential for the function of membrane proteins, current in vitro systems provide only limited options for studying them under biologically relevant conditions. Our experimental assay is based on the droplet-on-hydrogel bilayer technique (Leptihn et al. Nat Protoc 8:1048-1057, 2013), where a lipid bilayer forms between a sub-millimetre water droplet and a thin hydrogel layer on a glass cover slip, enabling high-resolution microscopy in total internal reflection mode. To extend the application of this assay beyond channels to other membrane proteins, we introduce a custom-built, electronically controlled perfusion system that is designed to directly connect to the droplet above the lipid bilayer. This system can supply a stable voltage to the bilayer and is suitable for delivery of fragile membrane proteins embedded in proteoliposomes via charged fusion (Ishmukhametov et al. Nat Commun 7:13025, 2016), introducing changes of chemical potentials, and timed introduction of labels or substrate into the droplet. This work represents one of the steps towards single-molecule functional study of F₁F_o ATP synthase under variable transmembrane potentials. High-resolution single-molecule observation of its rotation steps on the microsecond timescale could provide valuable insights into the mechanisms of energy transport across the molecule.

Supplementary information: The online version contains supplementary material available at 10.1007/s12551-025-01344-4.

The material properties of biomolecular condensates, such as interfacial tension, viscoelasticity, stiffness, and molecular dynamics, are crucial for their biological functions in processes like signal transduction, stress response, and gene regulation. These properties influence both endogenous condensates, like the nucleolus and stress granules, and synthetic condensates engineered for potential drug delivery applications. In vitro studies, using purified components, provide controlled environments to explore the fundamental physics of phase separation, offering high precision in manipulating molecular components and conditions. However, cell-based characterisations are indispensable for understanding the physiological relevance of biomolecular condensates, accounting for molecular crowding, post-translational modifications, and interactions with cellular structures. Light-microscopy techniques offer the potential to bridge in vitro findings with in cellulo behaviour. This review outlines some fundamental challenges of in cellulo studies and discusses the potential of fluorescently labelling biomolecular condensates using the tetracysteine tag/biarsenical dye strategy. We describe how fluorescence-based techniques, including fluorescence recovery after photobleaching (FRAP) and emerging techniques like fluorescence lifetime imaging microscopy (FLIM), flicker spectroscopy, and raster image correlation spectroscopy (RICS), may be used to gain a detailed understanding of the material properties of biomolecular condensates within the cellular environment. Finally, we discuss the potential of Brillouin light scattering (BLS) microscopy, a label-free technique that holds potential for deciphering the cellular biophysics of biomolecular condensates.

The paper describes approaches to coarse-grained modeling of mechanical movements of DNA. The theoretical results of applying the angular model to calculations of the natural oscillation frequency, the effect of medium viscosity on the probability of open states, and the distribution of mechanical energy in a DNA molecule under external influence are considered. Examples of the practical application of model calculations to assess the effect of disorders in the ATXN2 gene on the occurrence of additional zones of open states in the region of trinucleotide repeats are given. The practical application of coarse-grained models covers many areas: from fundamental studies of the mechanisms of DNA interaction with proteins to the development of new methods of genetic engineering and the creation of functional DNA nanomaterials. Coarse-grained DNA models demonstrate high versatility as a research tool in modern molecular biology and biophysics. Their fundamental advantage lies in the ability to effectively reproduce the key characteristics of molecular dynamics with significant savings in computing resources compared to full-atom modeling. The versatility of these models is manifested in their adaptability to the study of a wide range of biological phenomena. They allow one to adequately describe both local conformational changes in individual sections of a molecule and global mechanical properties of extended DNA structures. At the same time, the possibility of a detailed study of the interaction of a molecule with the environment, including the effect of solvent viscosity, is preserved. Of particular value is the ability to study both equilibrium and nonequilibrium processes over a wide range of time scales.

Calcium entry through voltage-gated calcium channels as well as calcium release from intracellular calcium depots is crucial for triggering the release of neurotransmitters, synaptic plasticity and regulating the activity of various intracellular proteins. Recent evidence suggests that neuronal calcium signaling is impaired in many neurodegenerative disorders. This review presents methodological approaches that allow the estimation of the dynamics of calcium entry in motor nerve endings using fluorescent calcium indicators. The methods considered were: selecting calcium dyes depending on the experimental conditions, and loading indicators into the cells of interest. This review outlines general principles and techniques to record fast fluorescent calcium signals. Possible results that can be obtained within the framework of experiments using the described methods are also presented.

Supplementary information: The online version contains supplementary material available at 10.1007/s12551-025-01336-4.

Structural biology has seen the evolution of multiple pioneering experimental techniques over the last few decades, with leaps in technology and software facilitating rapid solution of crystal structures and the 'resolution revolution' in cryo-electron microscopy. Higher magnetic field strengths have expanded the development of magnetic resonance techniques and their ability to study protein dynamics and conformational diversity. Moreover, decades of experimental data collection and public data deposition combined with modern machine-learning technology have now made it possible to computationally predict three-dimensional protein structures from their amino acid sequence within minutes using AlphaFold (AF), a feat that has inspired a new wave of research. AlphaFold now contributes towards experimental structure solution and provides plausible predictions for structured regions of proteins leaving dynamics and conformational exchange as the next major questions in the field. Nuclear magnetic resonance (NMR) spectroscopy is uniquely placed both to rapidly validate AF predictions and probe protein dynamics at an atomic level in solution. Electron paramagnetic resonance (EPR) spectroscopy can measure distances between specific points in large protein complexes and provide local and global ranges of movement. This review will explore the revival of magnetic resonance techniques in a post-AlphaFold landscape and address their importance in protein research.