Interest in HDAC3 inhibitors (HDAC3i) for pharmacological applications outside of cancer is growing. However, concerns regarding the possible mutagenicity of the commonly used hydroxamates (zinc-binding groups, ZBGs) are also increasing. Considering these concerns, non-hydroxamate ZBGs offer a promising alternative for the development of non-mutagenic HDAC3 inhibitors. Unfortunately, the quantum chemical space of non-hydroxamates has not been studied in detail. This study has three primary goals: (i) to perform semiempirical quantum chemical calculations, examining AM-1 model parameters relevant to zinc binding, (ii) to develop supervised mathematical learning models to train a diverse set of non-hydroxamate-based HDAC3i, and (iii) to apply fragment-based approaches to identify sub-structural fragments (fingerprints) that promote or hinder HDAC3 inhibitory activity through classification-based QSARs. In addition, flexible molecular docking analysis, 200 ns MD simulation, and free energy landscape (FEL) analysis further established the importance of identified fingerprints in the modulation of HDAC3 inhibitory activity. This comprehensive analysis of structural variations among non-hydroxamate HDAC3i provides valuable insights, contributing to the design of potential non-mutagenic HDAC3i.
{"title":"A semiempirical and machine learning approach for fragment-based structural analysis of non-hydroxamate HDAC3 inhibitors","authors":"Sk. Abdul Amin , Lucia Sessa , Rajdip Tarafdar , Shovanlal Gayen , Stefano Piotto","doi":"10.1016/j.bpc.2025.107409","DOIUrl":"10.1016/j.bpc.2025.107409","url":null,"abstract":"<div><div>Interest in HDAC3 inhibitors (HDAC3i) for pharmacological applications outside of cancer is growing. However, concerns regarding the possible mutagenicity of the commonly used hydroxamates (zinc-binding groups, ZBGs) are also increasing. Considering these concerns, non-hydroxamate ZBGs offer a promising alternative for the development of non-mutagenic HDAC3 inhibitors. Unfortunately, the quantum chemical space of non-hydroxamates has not been studied in detail. This study has three primary goals: (i) to perform semiempirical quantum chemical calculations, examining AM-1 model parameters relevant to zinc binding, (ii) to develop supervised mathematical learning models to train a diverse set of non-hydroxamate-based HDAC3i, and (iii) to apply fragment-based approaches to identify sub-structural fragments (fingerprints) that promote or hinder HDAC3 inhibitory activity through classification-based QSARs. In addition, flexible molecular docking analysis, 200 ns MD simulation, and free energy landscape (FEL) analysis further established the importance of identified fingerprints in the modulation of HDAC3 inhibitory activity. This comprehensive analysis of structural variations among non-hydroxamate HDAC3i provides valuable insights, contributing to the design of potential non-mutagenic HDAC3i.</div></div>","PeriodicalId":8979,"journal":{"name":"Biophysical chemistry","volume":"320 ","pages":"Article 107409"},"PeriodicalIF":3.3,"publicationDate":"2025-02-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143436748","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-02-15DOI: 10.1016/j.bpc.2025.107411
V.S. Manu , Giuseppe Melacini , Evgenii L. Kovrigin , J. Patrick Loria , Gianluigi Veglia
Biological macromolecules are dynamic entities that transition between various conformational states, often playing a vital role in biological functions. Their inherent flexibility spans a broad range of timescales. Motions occurring within the microsecond to millisecond range are especially important, as they are integral to processes such as enzyme catalysis, folding, ligand binding, and allostery. NMR Carr-Purcell-Meiboom-Gill (CPMG) relaxation dispersion measurements are the preferred method for characterizing macromolecular motions at atomic resolution. However, it is still uncertain whether the functional motions of multiple residues in macromolecules need to be coordinated and/or synchronized within the protein matrix in order to perform the desired function. Here, we illustrate an unbiased method to analyze NMR relaxation dispersion and identify dynamic clusters of residues that fluctuate on similar timescales within proteins. The method requires relaxation dispersion data for backbone amides or side-chain methyl groups, which are globally fitted using the Bloch-McConnell equations for each pair of residues. The goodness of the pairwise fitting serves as a metric to construct two-dimensional synchronous dynamics (SyncDyn) maps, allowing us to identify residue clusters whose dynamics are influenced by ligand binding. We applied our method to the catalytic subunit of the cAMP-dependent protein kinase A (PKAC) and the T17A mutant of ribonuclease A (RNAse A). The SyncDyn maps for PKA-C showed distinct clusters of residues located in critical allosteric sites. Nucleotide binding activates the movement of residues at the interface between the two lobes and also those distal to the active site. In the case of RNAse A, the SyncDyn maps show that residues fluctuating with the same time scale are interspersed in both lobes of the enzyme. Overall, our approach eliminates arbitrary manual selection of residues for dynamic clustering and objectively identifies all possible residue pairs that fluctuate synchronously, i.e. on the same timescale.
{"title":"Unbiased clustering of residues undergoing synchronous motions in proteins using NMR spin relaxation data","authors":"V.S. Manu , Giuseppe Melacini , Evgenii L. Kovrigin , J. Patrick Loria , Gianluigi Veglia","doi":"10.1016/j.bpc.2025.107411","DOIUrl":"10.1016/j.bpc.2025.107411","url":null,"abstract":"<div><div>Biological macromolecules are dynamic entities that transition between various conformational states, often playing a vital role in biological functions. Their inherent flexibility spans a broad range of timescales. Motions occurring within the microsecond to millisecond range are especially important, as they are integral to processes such as enzyme catalysis, folding, ligand binding, and allostery. NMR Carr-Purcell-Meiboom-Gill (CPMG) relaxation dispersion measurements are the preferred method for characterizing macromolecular motions at atomic resolution. However, it is still uncertain whether the functional motions of multiple residues in macromolecules need to be coordinated and/or synchronized within the protein matrix in order to perform the desired function. Here, we illustrate an unbiased method to analyze NMR relaxation dispersion and identify dynamic clusters of residues that fluctuate on similar timescales within proteins. The method requires relaxation dispersion data for backbone amides or side-chain methyl groups, which are globally fitted using the Bloch-McConnell equations for each pair of residues. The goodness of the pairwise fitting serves as a metric to construct two-dimensional synchronous dynamics (SyncDyn) maps, allowing us to identify residue clusters whose dynamics are influenced by ligand binding. We applied our method to the catalytic subunit of the cAMP-dependent protein kinase A (PKA<img>C) and the T17A mutant of ribonuclease A (RNAse A). The SyncDyn maps for PKA-C showed distinct clusters of residues located in critical allosteric sites. Nucleotide binding activates the movement of residues at the interface between the two lobes and also those distal to the active site. In the case of RNAse A, the SyncDyn maps show that residues fluctuating with the same time scale are interspersed in both lobes of the enzyme. Overall, our approach eliminates arbitrary manual selection of residues for dynamic clustering and objectively identifies all possible residue pairs that fluctuate synchronously, <em>i.e.</em> on the same timescale.</div></div>","PeriodicalId":8979,"journal":{"name":"Biophysical chemistry","volume":"320 ","pages":"Article 107411"},"PeriodicalIF":3.3,"publicationDate":"2025-02-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143444898","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-02-14DOI: 10.1016/j.bpc.2025.107410
Sara Abigail Ramírez-Cortés , Adrián Durán-Vargas , Jesús Antonio Rauda-Ceja , Paola Mendoza-Espinosa , Luis Fernando Cofas-Vargas , Armando Cruz-Rangel , Julio Isael Pérez-Carreón , Enrique García-Hernández
Prostaglandin reductase 1 (PTGR1) is an NADPH-dependent enzyme critical to eicosanoid metabolism. Its elevated expression in malignant tumors often correlates with poor prognosis due to its role in protecting cells against reactive oxygen species. This study explores the inhibitory potential of licochalcone A, a flavonoid derived from Xinjiang licorice root, on human PTGR1. Using molecular dynamics simulations, we mapped the enzyme's conformational landscape, revealing a low-energy, rigid-body-like movement of the catalytic domain relative to the nucleotide-binding domain that governs PTGR1's transition between open and closed states. Simulations of NADPH-depleted dimer and NADPH-bound monomer highlighted the critical role of intersubunit interactions and coenzyme binding in defining PTGR1's conformational landscape, offering a deeper understanding of its functional adaptability as a holo-homodimer. Covalent docking, informed by prior chemoproteomic cross-linking data, revealed a highly favorable binding pose for licochalcone A at the NADPH-binding site. This pose aligned with a transient noncovalent binding pose inferred from solvent site-guided molecular docking, emphasizing the stereochemical complementarity of the coenzyme-binding site to licochalcone A. Sequence analysis across PTGR1 orthologs in vertebrates and exploration of 3D structures of human NADPH-binding proteins further underscore the potential of the coenzyme-binding site as a scaffold for developing PTGR1-specific inhibitors, positioning licochalcone A as a promising lead compound.
{"title":"Targeting human prostaglandin reductase 1 with Licochalcone A: Insights from molecular dynamics and covalent docking studies","authors":"Sara Abigail Ramírez-Cortés , Adrián Durán-Vargas , Jesús Antonio Rauda-Ceja , Paola Mendoza-Espinosa , Luis Fernando Cofas-Vargas , Armando Cruz-Rangel , Julio Isael Pérez-Carreón , Enrique García-Hernández","doi":"10.1016/j.bpc.2025.107410","DOIUrl":"10.1016/j.bpc.2025.107410","url":null,"abstract":"<div><div>Prostaglandin reductase 1 (PTGR1) is an NADPH-dependent enzyme critical to eicosanoid metabolism. Its elevated expression in malignant tumors often correlates with poor prognosis due to its role in protecting cells against reactive oxygen species. This study explores the inhibitory potential of licochalcone A, a flavonoid derived from Xinjiang licorice root, on human PTGR1. Using molecular dynamics simulations, we mapped the enzyme's conformational landscape, revealing a low-energy, rigid-body-like movement of the catalytic domain relative to the nucleotide-binding domain that governs PTGR1's transition between open and closed states. Simulations of NADPH-depleted dimer and NADPH-bound monomer highlighted the critical role of intersubunit interactions and coenzyme binding in defining PTGR1's conformational landscape, offering a deeper understanding of its functional adaptability as a holo-homodimer. Covalent docking, informed by prior chemoproteomic cross-linking data, revealed a highly favorable binding pose for licochalcone A at the NADPH-binding site. This pose aligned with a transient noncovalent binding pose inferred from solvent site-guided molecular docking, emphasizing the stereochemical complementarity of the coenzyme-binding site to licochalcone A. Sequence analysis across PTGR1 orthologs in vertebrates and exploration of 3D structures of human NADPH-binding proteins further underscore the potential of the coenzyme-binding site as a scaffold for developing PTGR1-specific inhibitors, positioning licochalcone A as a promising lead compound.</div></div>","PeriodicalId":8979,"journal":{"name":"Biophysical chemistry","volume":"320 ","pages":"Article 107410"},"PeriodicalIF":3.3,"publicationDate":"2025-02-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143429134","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Phase segregation and domain formation in cell membranes and model lipid bilayers have become a relevant topic in the last decades due to their role in important cell functions such as signaling and molecule-membrane interactions. To date, the most accepted explanation for the formation of these domains in mammalian cells is that cholesterol-enriched sphingomyelin patches of membrane form because of the preferential interaction between them. However, detailed information on molecular interactions within cholesterol-containing bilayers and their comparison with other sterol-containing bilayers, such as those containing ergosterol, is needed to understand the role these molecules have. Recent experimental findings have shown sterol-dependent differences in the morphology of supported lipid bilayers, but the molecular basis for these differences remains unclear. This work provides a molecular explanation for these differences using atomistic Molecular Dynamics simulations of lipid bilayers composed of 1-palmitoyl-2-oleoyl-glycero-3-phosphocholine (POPC) and N-palmitoyl-D-erythro-sphingosylphosphorylcholine (PSM) with 20 mol% of cholesterol or ergosterol. Atomic force microscopy was used to validate the simulation. The simulation ran for 11 μs and revealed that both sterols affect the morphology of the membrane. Key findings include: ergosterol induces greater order in PSM domains compared to cholesterol, lipid diffusion constants are lower in ergosterol-containing membranes, sterol flip-flop rates are significantly reduced in ergosterol-containing membranes and ergosterol leads to greater PSM-sterol enrichment. These molecular-level differences provide insight into the experimentally observed variations in domain formation and membrane properties between cholesterol and ergosterol-containing bilayers. Our findings contribute to the understanding of sterol-specific effects on membrane organization and dynamics, with potential implications for cellular processes and drug interactions in different organisms.
Statement of significance
This study advances our understanding of how different sterols influence membrane properties through molecular dynamics simulations of three-component lipid membranes. Specifically, we investigate the effects of two major sterols: ergosterol, predominantly found in plants and fungi, and cholesterol, characteristic of mammalian cells. While extensive research has elucidated cholesterol's impact on lipid bilayers, studies on ergosterol's effects are comparatively limited. Our work provides a comprehensive comparison of these sterols, highlighting their similarities and differences. These insights not only enhance our knowledge of cell membrane structure and function, but also contribute to our understanding of selective drug permeability across membranes. This research has potential implications for both fundamental cell biology and pharmaceutical applications.
在过去的几十年里，细胞膜和模型脂质双层中的相分离和结构域形成已经成为一个相关的话题，因为它们在重要的细胞功能中起着重要的作用，如信号传导和分子-膜相互作用。迄今为止，对这些结构域在哺乳动物细胞中形成的最被接受的解释是，由于它们之间的优先相互作用，膜上富含胆固醇的鞘磷脂斑块形成。然而，要了解这些分子的作用，需要了解含胆固醇双分子层内部分子相互作用的详细信息，以及它们与其他含胆固醇双分子层（如含麦角甾醇的双分子层）的比较。最近的实验发现显示了甾醇依赖性的脂质双分子层形态差异，但这些差异的分子基础尚不清楚。本研究利用原子分子动力学模拟了由1-棕榈酰-2-油酰-甘油-3-磷酸胆碱（POPC）和n -棕榈酰- d -红-鞘酰基磷酸胆碱（PSM）与20 mol%的胆固醇或麦角甾醇组成的脂质双分子层，为这些差异提供了分子解释。采用原子力显微镜对模拟结果进行验证。模拟时间为11 μs，结果表明这两种甾醇都会影响膜的形态。主要发现包括：与胆固醇相比，麦角甾醇诱导PSM结构域的有序性更高，含麦角甾醇膜中的脂质扩散常数更低，含麦角甾醇膜中的甾醇转换率显著降低，麦角甾醇导致更大的PSM-甾醇富集。这些分子水平上的差异提供了对实验观察到的胆固醇和麦角甾醇双分子层之间结构域形成和膜性质的变化的见解。我们的发现有助于理解甾醇对膜组织和动力学的特异性作用，对不同生物体的细胞过程和药物相互作用具有潜在的影响。本研究通过对三组分脂质膜的分子动力学模拟，加深了我们对不同甾醇如何影响膜性质的理解。具体来说，我们研究了两种主要甾醇的作用：麦角甾醇，主要存在于植物和真菌中，以及胆固醇，哺乳动物细胞的特征。虽然广泛的研究已经阐明了胆固醇对脂质双分子层的影响，但对麦角甾醇作用的研究相对有限。我们的工作提供了这些固醇的全面比较，突出他们的异同。这些发现不仅增强了我们对细胞膜结构和功能的认识，而且有助于我们对药物选择性跨膜渗透的理解。这项研究对基础细胞生物学和药物应用都有潜在的意义。
{"title":"Effect of ergosterol or cholesterol on the morphology and dynamics of the POPC/sphingomyelin bilayer","authors":"Fernando Favela-Rosales , Jorge Hernández-Cobos , Arturo Galván-Hernández , Omar Hernández-Villanueva , Iván Ortega-Blake","doi":"10.1016/j.bpc.2025.107408","DOIUrl":"10.1016/j.bpc.2025.107408","url":null,"abstract":"<div><div>Phase segregation and domain formation in cell membranes and model lipid bilayers have become a relevant topic in the last decades due to their role in important cell functions such as signaling and molecule-membrane interactions. To date, the most accepted explanation for the formation of these domains in mammalian cells is that cholesterol-enriched sphingomyelin patches of membrane form because of the preferential interaction between them. However, detailed information on molecular interactions within cholesterol-containing bilayers and their comparison with other sterol-containing bilayers, such as those containing ergosterol, is needed to understand the role these molecules have. Recent experimental findings have shown sterol-dependent differences in the morphology of supported lipid bilayers, but the molecular basis for these differences remains unclear. This work provides a molecular explanation for these differences using atomistic Molecular Dynamics simulations of lipid bilayers composed of 1-palmitoyl-2-oleoyl-glycero-3-phosphocholine (POPC) and N-palmitoyl-D-erythro-sphingosylphosphorylcholine (PSM) with 20 mol% of cholesterol or ergosterol. Atomic force microscopy was used to validate the simulation. The simulation ran for 11 μs and revealed that both sterols affect the morphology of the membrane. Key findings include: ergosterol induces greater order in PSM domains compared to cholesterol, lipid diffusion constants are lower in ergosterol-containing membranes, sterol flip-flop rates are significantly reduced in ergosterol-containing membranes and ergosterol leads to greater PSM-sterol enrichment. These molecular-level differences provide insight into the experimentally observed variations in domain formation and membrane properties between cholesterol and ergosterol-containing bilayers. Our findings contribute to the understanding of sterol-specific effects on membrane organization and dynamics, with potential implications for cellular processes and drug interactions in different organisms.</div></div><div><h3>Statement of significance</h3><div>This study advances our understanding of how different sterols influence membrane properties through molecular dynamics simulations of three-component lipid membranes. Specifically, we investigate the effects of two major sterols: ergosterol, predominantly found in plants and fungi, and cholesterol, characteristic of mammalian cells. While extensive research has elucidated cholesterol's impact on lipid bilayers, studies on ergosterol's effects are comparatively limited. Our work provides a comprehensive comparison of these sterols, highlighting their similarities and differences. These insights not only enhance our knowledge of cell membrane structure and function, but also contribute to our understanding of selective drug permeability across membranes. This research has potential implications for both fundamental cell biology and pharmaceutical applications.</div></div>","PeriodicalId":8979,"journal":{"name":"Biophysical chemistry","volume":"320 ","pages":"Article 107408"},"PeriodicalIF":3.3,"publicationDate":"2025-02-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143444897","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-02-12DOI: 10.1016/j.bpc.2025.107412
Alexander S. Tatikolov , Pavel G. Pronkin , Ina G. Panova
Bilirubin, a product of heme catabolism, is toxic at elevated concentrations (>250–300 μM in blood serum), whereas at therapeutic concentrations (∼20–200 μM) exerts potent antioxidant, anti-inflammatory, immunomodulatory, cytoprotective and neuroprotective effects. Despite the therapeutic potential, its use in clinical practice is hampered by poor aqueous solubility, instability, and rapid metabolism. Nanotechnology overcomes these limitations and additionally imparts to bilirubin the advantages characteristic of nanopreparations: targeted action on the desired organ/tissue, increased therapeutic efficacy by delaying drug elimination from the body, improved transportation over biological barriers, the ability to combine therapeutic and diagnostic properties in a single agent. The review analyses the chemical synthesis, therapeutic mechanisms, and preclinical applications of nanosystems comprising bilirubin. In particular, nanostructures obtained by the covalent binding of bilirubin to macromolecules, bilirubin encapsulation in nanocarriers, bilirubin conjugation with metal nanoparticles and nanofunctionalization of inorganic compounds are considered; the data on the therapeutic trials of nanobilirubin are summarized. While studies on animal models and in vitro systems demonstrate improved biodistribution, reduced toxicity, and enhanced efficacy, no clinical trials to date have validated nanobilirubin formulations. Key barriers may include unresolved challenges in scalable synthesis, long-term biocompatibility, reproducible dosing of nanoformulations. Hence, further development of nanotherapeutic bilirubin agents for clinical practice is urgent.
{"title":"Bilirubin nanotechnology: An innovative approach in biomedicine","authors":"Alexander S. Tatikolov , Pavel G. Pronkin , Ina G. Panova","doi":"10.1016/j.bpc.2025.107412","DOIUrl":"10.1016/j.bpc.2025.107412","url":null,"abstract":"<div><div>Bilirubin, a product of heme catabolism, is toxic at elevated concentrations (>250–300 μM in blood serum), whereas at therapeutic concentrations (∼20–200 μM) exerts potent antioxidant, anti-inflammatory, immunomodulatory, cytoprotective and neuroprotective effects. Despite the therapeutic potential, its use in clinical practice is hampered by poor aqueous solubility, instability, and rapid metabolism. Nanotechnology overcomes these limitations and additionally imparts to bilirubin the advantages characteristic of nanopreparations: targeted action on the desired organ/tissue, increased therapeutic efficacy by delaying drug elimination from the body, improved transportation over biological barriers, the ability to combine therapeutic and diagnostic properties in a single agent. The review analyses the chemical synthesis, therapeutic mechanisms, and preclinical applications of nanosystems comprising bilirubin. In particular, nanostructures obtained by the covalent binding of bilirubin to macromolecules, bilirubin encapsulation in nanocarriers, bilirubin conjugation with metal nanoparticles and nanofunctionalization of inorganic compounds are considered; the data on the therapeutic trials of nanobilirubin are summarized. While studies on animal models and in vitro systems demonstrate improved biodistribution, reduced toxicity, and enhanced efficacy, no clinical trials to date have validated nanobilirubin formulations. Key barriers may include unresolved challenges in scalable synthesis, long-term biocompatibility, reproducible dosing of nanoformulations. Hence, further development of nanotherapeutic bilirubin agents for clinical practice is urgent.</div></div>","PeriodicalId":8979,"journal":{"name":"Biophysical chemistry","volume":"320 ","pages":"Article 107412"},"PeriodicalIF":3.3,"publicationDate":"2025-02-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143429135","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-02-03DOI: 10.1016/j.bpc.2025.107401
Celia Fricke , Jelica Milošević , Andreas Carlsson , Lars Boyens-Thiele , Marija Dubackic , Ulf Olsson , Alexander K. Buell , Sara Linse
The chaperone DNAJB6b (JB6) plays important roles in increasing amyloid protein solubility and inhibiting amyloid fibril formation, a causative factor for neurodegenerative diseases like Alzheimer's and Parkinson's disease. Insights into the biophysical properties of JB6, including its structure, self-assembly and stability towards denaturation, may enhance the understanding of the physicochemical basis of chaperone action. However, many of the biophysical properties of the chaperone remain elusive. Here, we investigated the structure and stability of JB6 and its domains towards thermal and chemical denaturation using Fourier transform infrared and circular dichroism spectroscopy to examine the thermodynamic properties. Both domains act as independent folding units. We find that the N-terminal domain (NTD) of JB6 is more stable than its C-terminal domain (CTD). Both domains are stabilized in the context of the full-length protein. The intact protein unfolds in a step-wise manner when subjected to a denaturing agent with the CTD unfolding at a lower denaturant concentration than the NTD. The combination of thermal and chemical denaturation followed by differential scanning fluorimetry revealed the enthalpy changes (22.6 and 26.4 kJ mol−1) and heat capacity changes (2.8 and 3.0 kJ/(mol*K)) upon denaturation of NTD alone and of NTD within the full-length protein, respectively. The understanding of JB6's biophysical properties complements the increasing amount of data on JB6's interactions with client proteins, paving the way for further investigation of the mechanism of its cellular function.
{"title":"On the thermal and chemical stability of DNAJB6b and its globular domains","authors":"Celia Fricke , Jelica Milošević , Andreas Carlsson , Lars Boyens-Thiele , Marija Dubackic , Ulf Olsson , Alexander K. Buell , Sara Linse","doi":"10.1016/j.bpc.2025.107401","DOIUrl":"10.1016/j.bpc.2025.107401","url":null,"abstract":"<div><div>The chaperone DNAJB6b (JB6) plays important roles in increasing amyloid protein solubility and inhibiting amyloid fibril formation, a causative factor for neurodegenerative diseases like Alzheimer's and Parkinson's disease. Insights into the biophysical properties of JB6, including its structure, self-assembly and stability towards denaturation, may enhance the understanding of the physicochemical basis of chaperone action. However, many of the biophysical properties of the chaperone remain elusive. Here, we investigated the structure and stability of JB6 and its domains towards thermal and chemical denaturation using Fourier transform infrared and circular dichroism spectroscopy to examine the thermodynamic properties. Both domains act as independent folding units. We find that the N-terminal domain (NTD) of JB6 is more stable than its C-terminal domain (CTD). Both domains are stabilized in the context of the full-length protein. The intact protein unfolds in a step-wise manner when subjected to a denaturing agent with the CTD unfolding at a lower denaturant concentration than the NTD. The combination of thermal and chemical denaturation followed by differential scanning fluorimetry revealed the enthalpy changes (22.6 and 26.4 kJ mol<sup>−1</sup>) and heat capacity changes (2.8 and 3.0 kJ/(mol*K)) upon denaturation of NTD alone and of NTD within the full-length protein, respectively. The understanding of JB6's biophysical properties complements the increasing amount of data on JB6's interactions with client proteins, paving the way for further investigation of the mechanism of its cellular function.</div></div>","PeriodicalId":8979,"journal":{"name":"Biophysical chemistry","volume":"320 ","pages":"Article 107401"},"PeriodicalIF":3.3,"publicationDate":"2025-02-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143379129","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Steroidal C7 alcohols and their esters are perspective agents in drug discovery. In addition, hydroxylation at C7 position could allow further modification of steroidal moiety. Such transformation is performed easily by the enzymes. Human steroid 7α-hydroxylases CYP7A1 and CYP7B1 are key enzymes taking part in the biotransformation of cholestanes, androstanes, pregnanes. In the article, we are focusing on the results of in vitro screening of a library of modified steroids toward CYP7 enzymes. A couple of compounds were found to express the affinity for binding to the enzymes, comparable with corresponding values for CYP7 natural ligands. Among them are 17-substituted androstane derivatives with N-containing pyridine ring and enone derivative of lithocholic acid, which bound by human CYP7A1, and D-seco and C16 oxime androstanes, which were identified as novel CYP7B1 ligands. Screening results revealed that both enzymes bind with high affinity a well-known drug abiraterone: in the case of CYP7A1 substrate-like binding mode was detected, with the formation of monohydroxylated product, while in case of CYP7B1 inhibitor-like binding was observed. Since CYP7 enzymes convert some of the studied compounds into their 7-hydroxy derivatives, potential of these enzymes as perspective regio- and stereoselective biocatalysts for obtaining C7 hydroxylated steroids could be assumed.
{"title":"Discovery of the potential of cholesterol-lowering human CYP7 enzymes as biocatalysts for the production of C7 hydroxylated steroids","authors":"Yaraslau Dzichenka , Michail Shapira , Antos Sachanka , Tatsiana Cherkesova , Veronika Shchur , Ljubica Grbović , Ksenija Pavlović , Bojana Vasiljević , Marina Savić , Andrea Nikolić , Aleksandar Oklješa , Jovana Ajduković , Ivana Kuzminac , Aliaksei Yantsevich , Sergey Usanov , Suzana Jovanović-Šanta","doi":"10.1016/j.bpc.2025.107393","DOIUrl":"10.1016/j.bpc.2025.107393","url":null,"abstract":"<div><div>Steroidal C7 alcohols and their esters are perspective agents in drug discovery. In addition, hydroxylation at C7 position could allow further modification of steroidal moiety. Such transformation is performed easily by the enzymes. Human steroid 7α-hydroxylases CYP7A1 and CYP7B1 are key enzymes taking part in the biotransformation of cholestanes, androstanes, pregnanes. In the article, we are focusing on the results of <em>in vitro</em> screening of a library of modified steroids toward CYP7 enzymes. A couple of compounds were found to express the affinity for binding to the enzymes, comparable with corresponding values for CYP7 natural ligands. Among them are 17-substituted androstane derivatives with N-containing pyridine ring and enone derivative of lithocholic acid, which bound by human CYP7A1, and D-seco and C16 oxime androstanes, which were identified as novel CYP7B1 ligands. Screening results revealed that both enzymes bind with high affinity a well-known drug abiraterone: in the case of CYP7A1 substrate-like binding mode was detected, with the formation of monohydroxylated product, while in case of CYP7B1 inhibitor-like binding was observed. Since CYP7 enzymes convert some of the studied compounds into their 7-hydroxy derivatives, potential of these enzymes as perspective regio- and stereoselective biocatalysts for obtaining C7 hydroxylated steroids could be assumed.</div></div>","PeriodicalId":8979,"journal":{"name":"Biophysical chemistry","volume":"319 ","pages":"Article 107393"},"PeriodicalIF":3.3,"publicationDate":"2025-01-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143144407","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Although radiation-induced clustered DNA damage can have critical biological consequences, the underlying molecular mechanisms remain unclear. To explore the effect of clustered DNA damage on DNA structure and dynamics, we performed molecular dynamics simulations on damaged DNA with two AP sites on the same strand, that is, a tandem AP cluster. The results showed that the cluster insertion of the two AP sites had a significant impact on the DNA's local and global structures. Local structural deformations as well as the extrahelical form, AP-base pairs, and irregular base pairs were frequently observed. Unlike a single AP site, the tandem AP cluster revealed that these local structural features occurred simultaneously within a small separation. Moreover, we found that the presence of tandem AP sites induced global bending of DNA. This suggests that the present case with tandem AP sites may have a non-negligible impact on the biological function of damage repair.
{"title":"Molecular dynamics study on clustered DNA damage: AP sites on the same strand","authors":"Kazushi Terakawa , Susumu Fujiwara , Tomoko Mizuguchi , Hiroaki Nakamura , Ken Akamatsu , Naoya Shikazono , Yoshiteru Yonetani","doi":"10.1016/j.bpc.2025.107394","DOIUrl":"10.1016/j.bpc.2025.107394","url":null,"abstract":"<div><div>Although radiation-induced clustered DNA damage can have critical biological consequences, the underlying molecular mechanisms remain unclear. To explore the effect of clustered DNA damage on DNA structure and dynamics, we performed molecular dynamics simulations on damaged DNA with two AP sites on the same strand, that is, a tandem AP cluster. The results showed that the cluster insertion of the two AP sites had a significant impact on the DNA's local and global structures. Local structural deformations as well as the extrahelical form, AP-base pairs, and irregular base pairs were frequently observed. Unlike a single AP site, the tandem AP cluster revealed that these local structural features occurred simultaneously within a small separation. Moreover, we found that the presence of tandem AP sites induced global bending of DNA. This suggests that the present case with tandem AP sites may have a non-negligible impact on the biological function of damage repair.</div></div>","PeriodicalId":8979,"journal":{"name":"Biophysical chemistry","volume":"319 ","pages":"Article 107394"},"PeriodicalIF":3.3,"publicationDate":"2025-01-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143073666","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-01-20DOI: 10.1016/j.bpc.2025.107392
Hagen Sülzen, Martin Klima, Vojtech Duchoslav, Evzen Boura
The development of small molecule drugs that target protein binders is the central goal in medicinal chemistry. During the lead compound development process, hundreds or even thousands of compounds are synthesized, with the primary focus on their binding affinity to protein targets. Typically, IC50 or EC50 values are used to rank these compounds. While thermodynamic values, such as the dissociation constant (KD), would be more informative, they are experimentally less accessible. In this study, we compare isothermal calorimetry (ITC) with surface plasmon resonance (SPR) using human STING, a key protein of innate immunity, and several cyclic dinucleotides (CDNs) that serve as its ligands. We demonstrate that SPR, with recent technological advancements, provides KDs that are sufficiently accurate for drug development purposes. To illustrate the versatility of our approach, we also used SPR to estimate the KD of poxin binding to cyclic GMP-AMP (cGAMP) that serves as a second messenger in the innate immune system. In conclusion, SPR offers a high benefit-to-cost ratio, making it an effective tool in the drug design process.
{"title":"SPR is a fast and straightforward method to estimate the binding constants of cyclic dinucleotides to their binding partners, such as STING or poxin","authors":"Hagen Sülzen, Martin Klima, Vojtech Duchoslav, Evzen Boura","doi":"10.1016/j.bpc.2025.107392","DOIUrl":"10.1016/j.bpc.2025.107392","url":null,"abstract":"<div><div>The development of small molecule drugs that target protein binders is the central goal in medicinal chemistry. During the lead compound development process, hundreds or even thousands of compounds are synthesized, with the primary focus on their binding affinity to protein targets. Typically, IC<sub>50</sub> or EC<sub>50</sub> values are used to rank these compounds. While thermodynamic values, such as the dissociation constant (KD), would be more informative, they are experimentally less accessible. In this study, we compare isothermal calorimetry (ITC) with surface plasmon resonance (SPR) using human STING, a key protein of innate immunity, and several cyclic dinucleotides (CDNs) that serve as its ligands. We demonstrate that SPR, with recent technological advancements, provides KDs that are sufficiently accurate for drug development purposes. To illustrate the versatility of our approach, we also used SPR to estimate the KD of poxin binding to cyclic GMP-AMP (cGAMP) that serves as a second messenger in the innate immune system. In conclusion, SPR offers a high benefit-to-cost ratio, making it an effective tool in the drug design process.</div></div>","PeriodicalId":8979,"journal":{"name":"Biophysical chemistry","volume":"319 ","pages":"Article 107392"},"PeriodicalIF":3.3,"publicationDate":"2025-01-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143027809","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-01-17DOI: 10.1016/j.bpc.2025.107389
Sayari Bhattacharya, Suman Chakrabarty
Quantitative characterization of protein conformational landscapes is a computationally challenging task due to their high dimensionality and inherent complexity. In this study, we systematically benchmark several widely used dimensionality reduction and clustering methods to analyze the conformational states of the Trp-Cage mini-protein, a model system with well-documented folding dynamics. Dimensionality reduction techniques, including Principal Component Analysis (PCA), Time-lagged Independent Component Analysis (TICA), and Variational Autoencoders (VAE), were employed to project the high-dimensional free energy landscape onto 2D spaces for visualization. Additionally, clustering methods such as K-means, hierarchical clustering, HDBSCAN, and Gaussian Mixture Models (GMM) were used to identify discrete conformational states directly in the high-dimensional space. Our findings reveal that density-based clustering approaches, particularly HDBSCAN, provide physically meaningful representations of free energy minima. While highlighting the strengths and limitations of each method, our study underscores that no single technique is universally optimal for capturing the complex folding pathways, emphasizing the necessity for careful selection and interpretation of computational methods in biomolecular simulations. These insights will contribute to refining the available tools for analyzing protein conformational landscapes, enabling a deeper understanding of folding mechanisms and intermediate states.
{"title":"Mapping conformational landscape in protein folding: Benchmarking dimensionality reduction and clustering techniques on the Trp-Cage mini-protein","authors":"Sayari Bhattacharya, Suman Chakrabarty","doi":"10.1016/j.bpc.2025.107389","DOIUrl":"10.1016/j.bpc.2025.107389","url":null,"abstract":"<div><div>Quantitative characterization of protein conformational landscapes is a computationally challenging task due to their high dimensionality and inherent complexity. In this study, we systematically benchmark several widely used dimensionality reduction and clustering methods to analyze the conformational states of the Trp-Cage mini-protein, a model system with well-documented folding dynamics. Dimensionality reduction techniques, including Principal Component Analysis (PCA), Time-lagged Independent Component Analysis (TICA), and Variational Autoencoders (VAE), were employed to project the high-dimensional free energy landscape onto 2D spaces for visualization. Additionally, clustering methods such as K-means, hierarchical clustering, HDBSCAN, and Gaussian Mixture Models (GMM) were used to identify discrete conformational states directly in the high-dimensional space. Our findings reveal that density-based clustering approaches, particularly HDBSCAN, provide physically meaningful representations of free energy minima. While highlighting the strengths and limitations of each method, our study underscores that no single technique is universally optimal for capturing the complex folding pathways, emphasizing the necessity for careful selection and interpretation of computational methods in biomolecular simulations. These insights will contribute to refining the available tools for analyzing protein conformational landscapes, enabling a deeper understanding of folding mechanisms and intermediate states.</div></div>","PeriodicalId":8979,"journal":{"name":"Biophysical chemistry","volume":"319 ","pages":"Article 107389"},"PeriodicalIF":3.3,"publicationDate":"2025-01-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143036684","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
扫码关注我们
求助内容：
应助结果提醒方式：
京公网安备 11010802042870号