Biophysica最新文献

{"title":"Molecular Survival Strategies Against Kidney Filtration: Implications for Therapeutic Protein Engineering.","authors":"William P Heaps, Anne Elise Packard, Kristina M McCammon, Tyler P Green, Joseph P Talley, Bradley C Bundy, Dennis Della Corte","doi":"10.3390/biophysica6010004","DOIUrl":"10.3390/biophysica6010004","url":null,"abstract":"<p><p>The glomerular filtration barrier poses a significant challenge for circulating proteins, with molecules below ~60-70 kDa facing rapid renal clearance. Endogenous proteins have evolved sophisticated evasion mechanisms including oligomerization, carrier binding, electrostatic repulsion, and FcRn-mediated recycling. Understanding these natural strategies provides blueprints for engineering therapeutic proteins with improved pharmacokinetics. This review examines how endogenous proteins resist filtration, evaluates their application in protein engineering, and discusses clinical translation including established technologies (PEGylation, Fc-fusion) and emerging strategies (albumin-binding domains, glycoengineering). We address critical challenges of balancing half-life extension with tissue penetration, biological activity, and immunogenicity-essential considerations for the rational design of next-generation therapeutics with optimized dosing and enhanced efficacy.</p>","PeriodicalId":72401,"journal":{"name":"Biophysica","volume":"6 1","pages":""},"PeriodicalIF":1.8,"publicationDate":"2026-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC13410728/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"148622772","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Integrating AI with Cellular and Mechanobiology: Trends and Perspectives","authors":"Sakib Mohammad, Md Sakhawat Hossain, Sydney L. Sarver","doi":"10.3390/biophysica5040062","DOIUrl":"https://doi.org/10.3390/biophysica5040062","url":null,"abstract":"Mechanobiology explores how physical forces and cellular mechanics influence biological processes. This field has experienced rapid growth, driven by advances in high-resolution imaging, micromechanical testing, and computational modeling. At the same time, the increasing complexity and volume of mechanobiological imaging and measurement data have made traditional analysis methods difficult to scale. Artificial intelligence (AI) has emerged as a practical tool to address these challenges by providing new methods for interpreting and predicting biological behavior. Recent studies have demonstrated potential in several areas, including image-based analysis of cell and nuclear morphology, traction force microscopy (TFM), cell segmentation, motility analysis, and the detection of cancer biomarkers. Within this context, we review AI applications that either incorporate mechanical inputs/outputs directly or infer mechanobiologically relevant information from cellular and nuclear structure. This study summarizes progress in four key domains: AI/ML-based cell morphology studies, cancer biomarker identification, cell segmentation, and prediction of traction forces and motility. We also discuss the advantages and limitations of integrating AI/ML into mechanobiological research. Finally, we highlight future directions, including physics-informed and hybrid AI approaches, multimodal data integration, generative strategies, and opportunities for computational biophysics-aligned applications.","PeriodicalId":72401,"journal":{"name":"Biophysica","volume":"5 4","pages":"62-62"},"PeriodicalIF":0.0,"publicationDate":"2025-12-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.mdpi.com/2673-4125/5/4/62/pdf?version=1765703443","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147921432","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Membrane Depth Measurements of E Protein by ²H ESEEM Spectroscopy in Lipid Bilayers.","authors":"Andrew K Morris, Robert M McCarrick, Gary A Lorigan","doi":"10.3390/biophysica5040058","DOIUrl":"10.3390/biophysica5040058","url":null,"abstract":"<p><p>A topological analysis was performed by taking ESEEM measurements of site-specifically labeled E protein from SARS-CoV-2. The intensity of deuterium modulation arising from either deuterated solvent or deuterated lipid acyl chains revealed exposure to solvent or the bilayer hydrophobic region. Spin-labeled lipids and soluble spin labels were used as points of comparison. The data indicate that spin labels placed along the transmembrane helix of the E protein showed close contact with lipid acyl chains, but also substantial contact with solvent, while those placed on the C-terminal domain showed substantial but lower exposure to lipid acyl chains, with comparable solvent exposure. The results support the view that the C-terminal domain is in contact with the bilayer surface.</p>","PeriodicalId":72401,"journal":{"name":"Biophysica","volume":"5 4","pages":""},"PeriodicalIF":1.8,"publicationDate":"2025-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12807508/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145999906","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Cytoskeletal Prestress Regulates RIG-I-Mediated Innate Immunity.","authors":"Arpan Roy, Sydney Sarver, Jarod Beights, Sean Brennan, Sazid Noor Rabi, Sakib Mohammad, Kyu Young Han, Sabrina Nilufar, Farhan Chowdhury","doi":"10.3390/biophysica5040051","DOIUrl":"10.3390/biophysica5040051","url":null,"abstract":"<p><p>Innate immunity is the body's first line of defense for mounting robust antiviral signaling. However, the role of cytoskeletal prestress, a hallmark of cellular mechanotransduction, in regulating innate immune pathways such as retinoic acid-inducible gene I (RIG-I) signaling remains poorly understood. Herein, we show that cells on soft vs. rigid substrates elicit cytoskeletal prestress-dependent activation of RIG-I signaling, leading to differential type-I interferon (IFN) gene expression. Cells were cultured on soft (0.6 kPa) and stiff (8.5 kPa) substrates to modulate cellular traction and prestress, followed by transfection of Poly(I:C), a synthetic viral dsRNA mimic, to measure the RIG-I-mediated innate immune response. Cells on soft substrates show minimal activation of RIG-I signaling, resulting in low expression of IFN-β1 and other IFN-stimulated genes (ISGs), compared to cells on stiff substrates. We further demonstrate that activation of TANK Binding Kinase 1 (TBK1), a downstream effector of the RIG-I pathway, is inhibited in cells on soft substrates due to the cytoplasmic sequestration of the Yes-associated protein (YAP), a HIPPO pathway effector protein. In contrast, cells on stiffer substrates experienced decreased TBK1 inhibition due to the nuclear localization of YAP and exhibited elevated TBK1 activation and heightened IFN and ISG expressions. Together, we demonstrate that cytoskeletal prestress represents a key biophysical regulator of innate immune signaling.</p>","PeriodicalId":72401,"journal":{"name":"Biophysica","volume":"5 4","pages":""},"PeriodicalIF":1.8,"publicationDate":"2025-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12889820/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"146168257","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Dynamic Features Control the Stabilization of the Green and Red Forms of the Chromophore in AzamiGreen Fluorescent Protein Variants","authors":"Vladimir B. Krapivin, Roman А. Stepanyuk, Maria G. Khrenova","doi":"10.3390/biophysica5040053","DOIUrl":"https://doi.org/10.3390/biophysica5040053","url":null,"abstract":"Fluorescent proteins find application as biocompatible, genetically encoded labels for visualization of living organisms tissues. Green fluorescent proteins (GFPs) are the most diverse, but proteins with red fluorescence have advantages, such as lower phototoxicity and better penetration into biological tissues. A promising approach is to obtain red fluorescent proteins (RFPs) from GFPs by introducing mutations that stabilize the oxidized chromophore state with an extended conjugated π-system. However, to date this remains a non-trivial task and experimental developments are carried out mainly by random mutagenesis. Development of descriptors obtained in molecular modeling can rationalize this field. Herein, we rely on experimental data on the AzamiGreen fluorescent protein and its variants that are oxidized to the red form. We perform classical molecular dynamics (MD) and combined quantum mechanics/molecular mechanics (QM/MM) simulations to determine structural and dynamic features that govern oxidation. We demonstrate that the red state is predominantly stabilized by interactions of polar lysine residues with chromophore oxygen atoms. Dynamic network analysis demonstrates that in red fluorescent proteins the chromophore motions are correlated with the movement of surrounding protein side chains to a higher extent than in green variants. The presence of different resonance forms of the chromophore determines the fluorescence band maximum value: a decrease in the phenolate form population leads to the red shift.","PeriodicalId":72401,"journal":{"name":"Biophysica","volume":"5 4","pages":"53-53"},"PeriodicalIF":0.0,"publicationDate":"2025-11-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.mdpi.com/2673-4125/5/4/53/pdf?version=1762789198","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147907680","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Isolation of an Anti-hG-CSF Nanobody and Its Application in Quantitation and Rapid Detection of hG-CSF in Pharmaceutical Testing","authors":"Qiang Ma, Liuqiang Zhu, Xiang Li, Dening Pei, Lei Yu, Xinchang Shi, Yong Zhou, Zhihao Fu, Chenggang Liang, Xi Qin, Junzhi Wang","doi":"10.3390/biophysica5040047","DOIUrl":"https://doi.org/10.3390/biophysica5040047","url":null,"abstract":"Human granulocyte colony-stimulating factor (hG-CSF) is primarily used to treat neutropenia induced by cancer chemotherapy and bone marrow transplantation. The current identification test for hG-CSF relies on Western blot (WB), a labor-intensive and technically demanding method. This study aimed to screen and prepare an anti-hG-CSF nanobody to identify and quantify hG-CSF, with the ultimate goal of developing colloidal gold-labeled nanobody test strips for rapid identification. An alpaca was immunized with hG-CSF, and the VHH gene sequence encoding the anti-hG-CSF nanobody was obtained through sequencing following phage display library construction and multiple rounds of biopanning. The nanobody C68, obtained from screening, was expressed by E. coli, and its physicochemical properties such as molecular weight, isoelectric point, and affinity were characterized after purification. WB analysis demonstrated excellent performance of the nanobody in identification tests in terms of specificity, limit of detection (LOD), applicability with products from various manufacturers, and thermal stability. Additionally, we established an ELISA method for hG-CSF quantification utilizing the nanobody C68 and conducted methodological validation. Finally, colloidal gold-based test strips were constructed using the nanobody C68, with a LOD of 30 μg/mL, achieving rapid identification for hG-CSF. This study represents a novel application of nanobodies in pharmaceutical testing and offers valuable insights for developing identification tests for other recombinant protein drugs.","PeriodicalId":72401,"journal":{"name":"Biophysica","volume":"5 4","pages":"47-47"},"PeriodicalIF":0.0,"publicationDate":"2025-10-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.mdpi.com/2673-4125/5/4/47/pdf?version=1760714172","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147902140","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Probing the Interaction Between Icariin and Proteinase K: A Combined Spectroscopic and Molecular Modeling Study","authors":"Zhongbao Han, Haoyuan Zheng, Yong Qi, Dilshadbek T. Usmanov, Liyan Liu, Zhan Yu","doi":"10.3390/biophysica5030032","DOIUrl":"https://doi.org/10.3390/biophysica5030032","url":null,"abstract":"Icariin (ICA) is widely recognized for its health benefits. In this work, we examined the intermolecular interactions between ICA and proteinase K (PK) via multi-spectroscopic techniques and molecular simulations. The experimental findings revealed that ICA quenched the fluorescence emission of PK by forming a noncovalent complex. Both hydrogen bonding and van der Waals interactions are essential for the complex’s formation. Then Förster resonance energy transfer (FRET), competitive experiments, and synchronous fluorescence spectroscopy were adopted to verify the formation of the complex. Molecular docking studies demonstrated that ICA could spontaneously bind to PK by hydrogen bonding and hydrophobic interactions, which is consistent with the spectroscopic results. The PK-ICA complex’s dynamic stability was evaluated using a 50 ns molecular dynamics (MD) simulation. The simulation results revealed no significant structural deformation or positional changes throughout the entire simulation period. The complex appears to be rather stable, as seen by the average root-mean-square deviation (RMSD) fluctuations for the host protein in the PK-ICA complex of 1.08 Å and 3.09 Å. These outcomes of molecular simulations suggest that ICA interacts spontaneously and tightly with PK, consistent with the spectroscopic findings. The approach employed in this research presents a pragmatic and advantageous method for examining protein–ligand interactions, as evidenced by the concordance between empirical and theoretical findings.","PeriodicalId":72401,"journal":{"name":"Biophysica","volume":"5 3","pages":"32-32"},"PeriodicalIF":0.0,"publicationDate":"2025-07-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.mdpi.com/2673-4125/5/3/32/pdf?version=1753691992","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"147332215","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Melanin in the Retinal Epithelium and Magnetic Sensing: A Review of Current Studies.","authors":"Lidia Zueva, Vassiliy Tsytsarev, Janaina Alves, Mikhail Inyushin","doi":"10.3390/biophysica4040030","DOIUrl":"10.3390/biophysica4040030","url":null,"abstract":"<p><p>Coming in a variety of forms, melanin is one of the most abundant, stable, diverse, and evolutionarily ancient pigments found in living things in nature. These pigments often serve protective functions, typically well-adapted to their specific roles. One such protective function is metal chelation and cation exchange, which help regulate and buffer metal concentrations within cells. By binding to certain metals, melanin can acquire magnetic properties. Because of this, it may play a role in magnetic effects and possibly in the response of organisms to external magnetic fields and magnetic sensing. While there is melanin in plants, microbes, fungi, and invertebrates, certain types of melanin are specifically associated with the retina in vertebrates, including migrating bird and fish species. In this review, we examine studies focusing on the properties of melanin in these parts of the body and their possible association with magnetic sensing, and generally, magnetic sensing in the retina.</p>","PeriodicalId":72401,"journal":{"name":"Biophysica","volume":"4 4","pages":"466-476"},"PeriodicalIF":0.0,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11500728/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142513973","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Anion Effect on Phase Separation of Polyethylene Glycol-8000–Sodium Salt Two-Phase Systems","authors":"Amber R. Titus, P. Madeira, V. Uversky, B. Zaslavsky","doi":"10.3390/biophysica4030025","DOIUrl":"https://doi.org/10.3390/biophysica4030025","url":null,"abstract":"Aqueous two-phase systems (ATPSs) are formed when two nonionic polymers, or a single polymer and salt, are mixed in water above a specific concentration, resulting in the emergence of phase separation and the formation of two immiscible aqueous phases. The solvent properties of the aqueous media within the phases of ATPSs rely on the specific composition of the co-solutes and the arrangement of the hydrogen bond network within each phase. Here, we investigate the anion effect of various sodium salts on the enhancement or destabilization of polyethylene glycol (PEG)–salt ATPS formation. Relatively small changes in ATPS ionic composition were shown to result in significant changes in solute partitioning. Additionally, we previously established that the arrangement of hydrogen bonds within the coexisting phases of ATPSs is different, as evidenced by Attenuated Total Reflection—Fourier Transform Infrared (ATR-FTIR) spectroscopic analysis of OH-stretch bands. The hydrogen bond arrangement was shown to abruptly change at concentrations below the threshold of macroscopic phase separation in the ATPSs. Using dynamic light scattering (DLS), we observed a correlation between these abrupt changes in H-bond arrangement and the detection of agglomerate formation in both polymer–polymer and polymer–salt systems.","PeriodicalId":72401,"journal":{"name":"Biophysica","volume":"43 23","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-07-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141654995","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Intermolecular FRET Pairs as An Approach to Visualize Specific Enzyme Activity in Model Biomembranes and Living Cells","authors":"I. D. Zlotnikov, Alexander A. Ezhov, E. V. Kudryashova","doi":"10.3390/biophysica4030024","DOIUrl":"https://doi.org/10.3390/biophysica4030024","url":null,"abstract":"Herein, we propose an analytical approach based on intermolecular fluorescent resonant energy transfer (FRET) pairs for the visualization of specific enzyme activity in model biomembranes and in living cells. Cell visualizations with fluorescent confocal laser microscopy usually rely on fluorescent probes, such as Fluorescein isothiocyanate (FITC), Alexa488, Tetramethylrhodamine isothiocyanate (TRITC) and many others. However, for more specific tasks, such as the detection of certain enzymatic activity inside the living cell, the toolbox is quite limited. In the case of enzyme-hydrolases for example, the choice is limited to organic molecules comprising a fluorescent dye (typically, 4-methylumbelliferone (MUmb) or 7-amino-4-methylcoumarin (AMC) derivatives) and a fluorescence quencher, bound via an enzyme-sensitive linker—so that when the linker is degraded, the fluorescent signal increases. Unfortunately, both MUmb and AMC are quenched and have a relatively low quantum yield in cells, and their excitation and emission ranges overlap with that of intracellular fluorophores, often producing a strong background noise. R6G, on the other hand, has excellent quantum yield apart from intracellular fluorophores, but there are no efficient quenchers that could be chemically linked to R6G. Herein, we show that R6G is able to form intermolecular FRET pairs with MUmb or AMC, with the latter serving as fluorescence donors. This yields a combination of R6G’s excellent fluorescence properties with a possibility to use an enzyme-sensitive linker in MUTMAC or AMC derivatives. This phenomenon was initially discovered in a model system, reversed micelles, where the donor, the acceptor, and the enzyme are forced to be in close proximity to each other, so that proximity could serve as an explanation for the intermolecular FRET effect. Surprisingly enough, the phenomenon has been reproduced in living cells. Moreover, we were able to create working intermolecular donor–acceptor FRET pairs for several different enzymes, including chymotrypsin, phosphatase, and asparaginase. This appears counterintuitive, as besides the overlap of the emission spectra of the donor and the absorption spectra of the acceptor, there are other criteria for the FRET effect, including the convergence of two fluorophores at a distance of about 1–10 nm, and the orientation of their dipoles at a certain angle, which is difficult to imagine in a bulk system like a living cell. We hypothesize that FRET-enabling donor–acceptor interaction may be taking place at the inner surface of the lipid bilayer, to which both donor and acceptor molecules would likely have an affinity. This hypothesis would require a more detailed investigation. Therefore, we have shown that the method suggested has good potential in the visualization of enzyme functioning inside living cells, which is often a challenging task. Shifting of the fluorescence signal to the long-wavelength region would increase the signal selectivi","PeriodicalId":72401,"journal":{"name":"Biophysica","volume":"12 2","pages":""},"PeriodicalIF":0.0,"publicationDate":"2024-07-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141704417","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}