Pub Date : 2025-12-01DOI: 10.1016/j.biochi.2025.09.008
Zimeng Han , Xuelian Han , Wanting Cheng , Shaolei Geng , Weidong Li , Tianyun Wang , Xiaoyin Wang
Mammalian cells are the primary host for recombinant protein production, and the vector system significantly impacts expression efficiency and stability. Traditional vectors often suffer from transgene silencing and clone heterogeneity, requiring large-scale screening to obtain stable and high-yield cell lines, which is time-consuming and labor-intensive. Transposon expression vector mediate highly efficient, semi-targeted integration of the gene of interest in mammalian cells. Integration occurs at transcriptionally active genomic loci of the host cell, thereby ensuring persistent expression. Moreover, non-viral transposon expression vectors offer a safer and more attractive alternative to viral vectors by avoiding potential tumorigenicity and immune reactions associated with viral proteins and oncogenes, making them ideal for gene therapy applications. This review summarizes the structure, mechanism, and optimization of transposon vectors, as well as their applications in recombinant protein expression and gene therapy.
{"title":"Application of transposon expression system in recombinant protein expression and gene therapy","authors":"Zimeng Han , Xuelian Han , Wanting Cheng , Shaolei Geng , Weidong Li , Tianyun Wang , Xiaoyin Wang","doi":"10.1016/j.biochi.2025.09.008","DOIUrl":"10.1016/j.biochi.2025.09.008","url":null,"abstract":"<div><div>Mammalian cells are the primary host for recombinant protein production, and the vector system significantly impacts expression efficiency and stability. Traditional vectors often suffer from transgene silencing and clone heterogeneity, requiring large-scale screening to obtain stable and high-yield cell lines, which is time-consuming and labor-intensive. Transposon expression vector mediate highly efficient, semi-targeted integration of the gene of interest in mammalian cells. Integration occurs at transcriptionally active genomic loci of the host cell, thereby ensuring persistent expression. Moreover, non-viral transposon expression vectors offer a safer and more attractive alternative to viral vectors by avoiding potential tumorigenicity and immune reactions associated with viral proteins and oncogenes, making them ideal for gene therapy applications. This review summarizes the structure, mechanism, and optimization of transposon vectors, as well as their applications in recombinant protein expression and gene therapy.</div></div>","PeriodicalId":251,"journal":{"name":"Biochimie","volume":"239 ","pages":"Pages 97-108"},"PeriodicalIF":3.0,"publicationDate":"2025-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145088177","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 : 2025-12-01DOI: 10.1016/j.biochi.2025.09.006
Jiye Fu , Tianyu Chen , Na Lu , Xuan Pan , Jing Tu
Guanine-rich sequences are widely distributed throughout the human genome and are capable of forming intramolecular G-quadruplex (G4) structures through Hoogsteen hydrogen bonding. These structures have been implicated in diverse regulatory processes. While extensive studies have established that loop architecture—particularly loop length and composition—profoundly affects G4 structural stability, most investigations have relied on synthetic sequences with predefined loop configurations that do not accurately reflect genomic contexts. In the current study, we analyzed the chain composition and stability of G-quadruplexes within the human genome to clarify the relationship between them by high throughput sequencing data. We utilized G4-forming sequences identified by G4-seq and G4-miner—two sequencing-based methods that detect G4s through polymerase stalling–associated drops in sequencing quality scores, where more stable structures produce stronger signals and thus higher detection rates—as the primary dataset. Our analysis revealed a negative correlation between total loop length and G4 stability, whereas individual loop length distributions exhibited minimal influence. Interestingly, G4s with short loops frequently occur in the genome as microsatellites or tandem atypical G4 arrays, resulting in structural stability profiles that deviate from those observed in synthetic G4 motifs in vitro. Molecular dynamics simulations incorporating native flanking sequences further corroborated these findings, underscoring the importance of genomic context in determining G4 stability. We note that the research was restricted to canonical G4s, which may limit the generality of our conclusions.
{"title":"Decoding G-quadruplex stability: the role of loop architecture and sequence context in the human genome","authors":"Jiye Fu , Tianyu Chen , Na Lu , Xuan Pan , Jing Tu","doi":"10.1016/j.biochi.2025.09.006","DOIUrl":"10.1016/j.biochi.2025.09.006","url":null,"abstract":"<div><div>Guanine-rich sequences are widely distributed throughout the human genome and are capable of forming intramolecular G-quadruplex (G4) structures through Hoogsteen hydrogen bonding. These structures have been implicated in diverse regulatory processes. While extensive studies have established that loop architecture—particularly loop length and composition—profoundly affects G4 structural stability, most investigations have relied on synthetic sequences with predefined loop configurations that do not accurately reflect genomic contexts. In the current study, we analyzed the chain composition and stability of G-quadruplexes within the human genome to clarify the relationship between them by high throughput sequencing data. We utilized G4-forming sequences identified by G4-seq and G4-miner—two sequencing-based methods that detect G4s through polymerase stalling–associated drops in sequencing quality scores, where more stable structures produce stronger signals and thus higher detection rates—as the primary dataset. Our analysis revealed a negative correlation between total loop length and G4 stability, whereas individual loop length distributions exhibited minimal influence. Interestingly, G4s with short loops frequently occur in the genome as microsatellites or tandem atypical G4 arrays, resulting in structural stability profiles that deviate from those observed in synthetic G4 motifs in vitro. Molecular dynamics simulations incorporating native flanking sequences further corroborated these findings, underscoring the importance of genomic context in determining G4 stability. We note that the research was restricted to canonical G4s, which may limit the generality of our conclusions.</div></div>","PeriodicalId":251,"journal":{"name":"Biochimie","volume":"239 ","pages":"Pages 236-243"},"PeriodicalIF":3.0,"publicationDate":"2025-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145058804","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 : 2025-12-01DOI: 10.1016/j.biochi.2025.10.007
Ravy Leon Foun Lin, Adam Bellaiche, Catherine Etchebest
Despite its relevance, the well-known paradigm that connects sequence, structure and function still overlooks an important factor: the dynamics of the proteins involved in functional mechanisms. Experimental techniques such as nuclear magnetic resonance and, more recently, cryo-electron microscopy, provide some insight into the conformational diversity of a protein. However, technical difficulties limit their application to the proteome scale. Nevertheless, computational methods are now considered efficient in providing valuable information about the dynamical landscape of a given protein, thereby improving our understanding of its function. Among these methods, molecular dynamics simulations have become very popular and generate a large amount of data that can now be used by Artificial Intelligence approaches for prediction. This paper will describe and discuss the concepts and a few applications of these approaches.
{"title":"The key role of the dynamics and flexibility of proteins in functional mechanisms: How computational methods can contribute to their identification","authors":"Ravy Leon Foun Lin, Adam Bellaiche, Catherine Etchebest","doi":"10.1016/j.biochi.2025.10.007","DOIUrl":"10.1016/j.biochi.2025.10.007","url":null,"abstract":"<div><div>Despite its relevance, the well-known paradigm that connects sequence, structure and function still overlooks an important factor: the dynamics of the proteins involved in functional mechanisms. Experimental techniques such as nuclear magnetic resonance and, more recently, cryo-electron microscopy, provide some insight into the conformational diversity of a protein. However, technical difficulties limit their application to the proteome scale. Nevertheless, computational methods are now considered efficient in providing valuable information about the dynamical landscape of a given protein, thereby improving our understanding of its function. Among these methods, molecular dynamics simulations have become very popular and generate a large amount of data that can now be used by Artificial Intelligence approaches for prediction. This paper will describe and discuss the concepts and a few applications of these approaches.</div></div>","PeriodicalId":251,"journal":{"name":"Biochimie","volume":"239 ","pages":"Pages 8-26"},"PeriodicalIF":3.0,"publicationDate":"2025-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145310230","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 : 2025-11-29DOI: 10.1016/j.biochi.2025.11.012
Chia-Chi Chang , Govindan Sivakumar , Meng-Chieh Huang , Yun-Hsuan Chen , Shan-Rong Wu , Yun-Ju Lai , Chian-Hui Lai , Ling-Hsien Tu
Human calcitonin (hCT) is a peptide hormone made up of 32 amino acids, produced by the parafollicular cells of the thyroid gland. Its main role is to decrease blood calcium levels by inhibiting osteoclast activity and reducing calcium reabsorption in the kidneys and intestines. This characteristic positions hCT as a potential treatment for bone-related conditions, such as osteoporosis and Paget's disease. However, hCT has a strong tendency to form amyloid fibrils, which can lead to a loss of its biological function. This research aims to stabilize hCT's conformation through molecular conjugation to prevent its aggregation. We used a cross-linking tool with a boronic ester structure (NPC) to react with the side-chain amino groups of two residues in hCT, effectively "locking" its structure. hCT can be "unlocked" using hydrogen peroxide, returning it to its original form. To support this experimental design, we synthesized two hCT variants, hCT-Q14K and hCT-F22K. Our results showed that both variants successfully conjugated with NPC, and the conjugation could be easily removed with hydrogen peroxide within 30 min. NPC-conjugated hCT did not form amyloid as revealed by transmission electron microscopy. Moreover, thioflavin-T fluorescence kinetic studies and dynamic light scattering measurement demonstrated that the aggregation tendencies of hCT-F22K were significantly lower than those of hCT, while still maintaining their biological activity, suggesting this design is feasible. This approach may significantly aid in the development of calcitonin drugs.
{"title":"Both single mutation and molecular conjugation on the mutant prevent calcitonin from forming amyloid fibrils","authors":"Chia-Chi Chang , Govindan Sivakumar , Meng-Chieh Huang , Yun-Hsuan Chen , Shan-Rong Wu , Yun-Ju Lai , Chian-Hui Lai , Ling-Hsien Tu","doi":"10.1016/j.biochi.2025.11.012","DOIUrl":"10.1016/j.biochi.2025.11.012","url":null,"abstract":"<div><div>Human calcitonin (hCT) is a peptide hormone made up of 32 amino acids, produced by the parafollicular cells of the thyroid gland. Its main role is to decrease blood calcium levels by inhibiting osteoclast activity and reducing calcium reabsorption in the kidneys and intestines. This characteristic positions hCT as a potential treatment for bone-related conditions, such as osteoporosis and Paget's disease. However, hCT has a strong tendency to form amyloid fibrils, which can lead to a loss of its biological function. This research aims to stabilize hCT's conformation through molecular conjugation to prevent its aggregation. We used a cross-linking tool with a boronic ester structure (<strong>NPC</strong>) to react with the side-chain amino groups of two residues in hCT, effectively \"locking\" its structure. hCT can be \"unlocked\" using hydrogen peroxide, returning it to its original form. To support this experimental design, we synthesized two hCT variants, hCT-Q14K and hCT-F22K. Our results showed that both variants successfully conjugated with <strong>NPC</strong>, and the conjugation could be easily removed with hydrogen peroxide within 30 min. <strong>NPC</strong>-conjugated hCT did not form amyloid as revealed by transmission electron microscopy. Moreover, thioflavin-T fluorescence kinetic studies and dynamic light scattering measurement demonstrated that the aggregation tendencies of hCT-F22K were significantly lower than those of hCT, while still maintaining their biological activity, suggesting this design is feasible. This approach may significantly aid in the development of calcitonin drugs.</div></div>","PeriodicalId":251,"journal":{"name":"Biochimie","volume":"242 ","pages":"Pages 1-8"},"PeriodicalIF":3.0,"publicationDate":"2025-11-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145650371","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 : 2025-11-22DOI: 10.1016/j.biochi.2025.11.011
Garry W. Buchko , Lijun Liu , Kevin P. Battaile , Justin K. Craig , Elizabeth K. Harmon , Wesley C. Van Voorhis , Peter J. Myler , Scott Lovell
C2 domains are ubiquitous membrane-binding modules of ∼130 residues in eukaryotes that are often associated with proteins involved in membrane trafficking and lipid modification. The genome of Trichomonas vaginalis, the most common, non-viral, sexually transmitted human pathogen, encodes eight genes that contain a N-terminal C2 module linked to a XYPPX-repeat domain of more than four XYPPX repeats (C2-XYPPX). While the function of the XYPPX-repeat domain remains unknown, its multiple association with C2 domains in T. vaginalis suggests it is important. The C2 domain from one of these C2-XYPPX-repeat proteins, Tv-C2-1, was structurally and physically characterized using X-ray crystallography and NMR spectroscopy. The crystal structure for Tv-C2-1 shows that this domain shares a fold common to all C2 domains, a compact Greek-key motif composed of eight anti-parallel β-strands in the type-2 topology. An NMR chemical shift perturbation study with Ca2+ showed that Tv-C2-1 bound two Ca2+ atoms primarily via two loops (loop-1 and loop-3) on the predicted calcium binding face of the protein with Kds of 58.0 ± 0.1 μM and 232 ± 6 μM. Estimations of the overall rotational correlation time, τc, in the apo (11.1 ns) and Ca2+-bound (9.2 ns) state suggests the protein becomes more compact upon Ca2+ binding, consistent with a decrease in dynamics in loop-3 and marginally in loop-1 suggested by amide 15N heteronuclear steady-state {1H}-15N NOEs. Showing Tv-C2-1 binds calcium and adopts a compact Greek-key motif structure, two primary features of C2 domains, suggests understanding the function of the XYPPX-repeat domain may be warranted.
{"title":"A Trichomonas vaginalis C2-XYPPX-repeat protein with a structured C2 domain displaying dampened flexibility upon binding calcium","authors":"Garry W. Buchko , Lijun Liu , Kevin P. Battaile , Justin K. Craig , Elizabeth K. Harmon , Wesley C. Van Voorhis , Peter J. Myler , Scott Lovell","doi":"10.1016/j.biochi.2025.11.011","DOIUrl":"10.1016/j.biochi.2025.11.011","url":null,"abstract":"<div><div>C2 domains are ubiquitous membrane-binding modules of ∼130 residues in eukaryotes that are often associated with proteins involved in membrane trafficking and lipid modification. The genome of <em>Trichomonas vaginalis,</em> the most common, non-viral, sexually transmitted human pathogen, encodes eight genes that contain a N-terminal C2 module linked to a XYPPX-repeat domain of more than four XYPPX repeats (C2-XYPPX). While the function of the XYPPX-repeat domain remains unknown, its multiple association with C2 domains in <em>T. vaginalis</em> suggests it is important. The C2 domain from one of these C2-XYPPX-repeat proteins, <em>Tv</em>-C2-1, was structurally and physically characterized using X-ray crystallography and NMR spectroscopy. The crystal structure for <em>Tv</em>-C2-1 shows that this domain shares a fold common to all C2 domains, a compact Greek-key motif composed of eight anti-parallel β-strands in the type-2 topology. An NMR chemical shift perturbation study with Ca<sup>2+</sup> showed that <em>Tv</em>-C2-1 bound two Ca<sup>2+</sup> atoms primarily via two loops (loop-1 and loop-3) on the predicted calcium binding face of the protein with K<sub>d</sub>s of 58.0 ± 0.1 μM and 232 ± 6 μM. Estimations of the overall rotational correlation time, τ<sub>c</sub>, in the apo (11.1 ns) and Ca<sup>2+</sup>-bound (9.2 ns) state suggests the protein becomes more compact upon Ca<sup>2+</sup> binding, consistent with a decrease in dynamics in loop-3 and marginally in loop-1 suggested by amide <sup>15</sup>N heteronuclear steady-state {<sup>1</sup>H}-<sup>15</sup>N NOEs. Showing <em>Tv</em>-C2-1 binds calcium and adopts a compact Greek-key motif structure, two primary features of C2 domains, suggests understanding the function of the XYPPX-repeat domain may be warranted.</div></div>","PeriodicalId":251,"journal":{"name":"Biochimie","volume":"241 ","pages":"Pages 44-55"},"PeriodicalIF":3.0,"publicationDate":"2025-11-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145597251","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 : 2025-11-19DOI: 10.1016/j.biochi.2025.11.005
Pegah Moshtaghian , Hoda Ayat , Ali Mohammad Ahadi , Hamzeh Oraie
Snake venom is a rich source of bioactive peptides with diverse pharmacological effects. Kunitz-type protease inhibitors are one of the multifunctional peptides isolated from the snake venom. The Razi's viper, Macrovipera razii, is a species endemic to Iran whose venom components are still largely unexplored. In this study, we report the isolation, cloning, and functional characterization of PIMR, a novel Kunitz-type protease inhibitor derived from M. razii venom. The gene encoding PIMR was amplified from venom gland cDNA and then cloned into a prokaryotic expression vector. The PIMR protein consists of 71 residues stabilized by three disulfide bonds, characteristic of the folded Kunitz domain. Recombinant PIMR was expressed in Escherichia coli and purified by affinity chromatography. The anticoagulant activity of PIMR was assessed by clotting time, prothrombin time, and activated partial thromboplastin time assays. The results indicated a dose-dependent anticoagulant activity of the peptide. The PIMR also exhibited anticancer effects against the highly invasive breast cancer cell line, MDA-MB-231, with an IC50 of 33 μg/μl. Docking studies indicated that the RGN motif of PIMR interacts with integrins, suggesting a potential mechanism for inhibiting cancer cell proliferation. Additionally, molecular docking confirmed binding of PIMR to trypsin, supporting its role as a serine protease inhibitor. Our findings establish PIMR as a novel serine protease inhibitor with promising anti-coagulant and anti-cancer properties. PIMR, with its diverse biological functions, represents a potential candidate for drug development and further molecular investigations.
{"title":"PIMR, a novel Kunitz-type protease inhibitor from the Macrovipera razii, with dual anticoagulant and anticancer activities","authors":"Pegah Moshtaghian , Hoda Ayat , Ali Mohammad Ahadi , Hamzeh Oraie","doi":"10.1016/j.biochi.2025.11.005","DOIUrl":"10.1016/j.biochi.2025.11.005","url":null,"abstract":"<div><div>Snake venom is a rich source of bioactive peptides with diverse pharmacological effects. Kunitz-type protease inhibitors are one of the multifunctional peptides isolated from the snake venom. The Razi's viper, <em>Macrovipera razii</em>, is a species endemic to Iran whose venom components are still largely unexplored. In this study, we report the isolation, cloning, and functional characterization of PIMR, a novel Kunitz-type protease inhibitor derived from <em>M. razii</em> venom. The gene encoding PIMR was amplified from venom gland cDNA and then cloned into a prokaryotic expression vector. The PIMR protein consists of 71 residues stabilized by three disulfide bonds, characteristic of the folded Kunitz domain. Recombinant PIMR was expressed in <em>Escherichia coli</em> and purified by affinity chromatography. The anticoagulant activity of PIMR was assessed by clotting time, prothrombin time, and activated partial thromboplastin time assays. The results indicated a dose-dependent anticoagulant activity of the peptide. The PIMR also exhibited anticancer effects against the highly invasive breast cancer cell line, MDA-MB-231, with an IC50 of 33 μg/μl. Docking studies indicated that the RGN motif of PIMR interacts with integrins, suggesting a potential mechanism for inhibiting cancer cell proliferation. Additionally, molecular docking confirmed binding of PIMR to trypsin, supporting its role as a serine protease inhibitor. Our findings establish PIMR as a novel serine protease inhibitor with promising anti-coagulant and anti-cancer properties. PIMR, with its diverse biological functions, represents a potential candidate for drug development and further molecular investigations.</div></div>","PeriodicalId":251,"journal":{"name":"Biochimie","volume":"241 ","pages":"Pages 106-115"},"PeriodicalIF":3.0,"publicationDate":"2025-11-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145575155","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}
Sodium benzoate (SB), a widely used food preservative, has been reported to possess several therapeutic benefits including treating neurodegenerative disorders as well as negative effects such as altering glucose homeostasis. However, limited attention has been given to its role in inducing insulin resistance (IR). Therefore, current study was designed to investigate the relationship between SB and insulin resistance using C2C12 myotubes and to explore the possible underlying mechanisms. Myotubes were exposed to SB (20 mM) for 24 h and glucose consumption and uptake assays along with confocal microscopy (GLUT4 translocation) were used to evaluate IR. Oxidative stress indicators i.e. reactive oxygen species (ROS), lipid peroxidation (LPO), and reduced glutathione (GSH) levels were also measured. Myotubes morphology along with atrophic markers (calpain activity) were evaluated. Data show that in C2C12 myotubes, SB treatment reduced glucose consumption and uptake in a dose-dependent manner. SB inducing IR by disrupting insulin-mediated GLUT4 translocation in cultured myotubes. Alteration in oxidative stress-related markers (i.e. elevated ROS and LPO levels, reduced GSH) were also observed in SB-treated myotubes. Furthermore, decrease in the fusion index, length, and diameter of myotubes along with upregulation of calpain activity and decrease in muscle protein were also observed. Study concludes that SB exposure not only induced IR but also caused atrophy and oxidative stress in the C2C12 myotubes.
{"title":"Sodium benzoate, a common food preservative-induced Insulin resistance and atrophy in C2C12 myotubes","authors":"Onkar Sharma , Anita Dua , Sanjeev Gupta , Elisha Injeti , Ashwani Mittal","doi":"10.1016/j.biochi.2025.11.009","DOIUrl":"10.1016/j.biochi.2025.11.009","url":null,"abstract":"<div><div>Sodium benzoate (SB), a widely used food preservative, has been reported to possess several therapeutic benefits including treating neurodegenerative disorders as well as negative effects such as altering glucose homeostasis. However, limited attention has been given to its role in inducing insulin resistance (IR). Therefore, current study was designed to investigate the relationship between SB and insulin resistance using C2C12 myotubes and to explore the possible underlying mechanisms. Myotubes were exposed to SB (20 mM) for 24 h and glucose consumption and uptake assays along with confocal microscopy (GLUT4 translocation) were used to evaluate IR. Oxidative stress indicators <em>i.e.</em> reactive oxygen species (ROS), lipid peroxidation (LPO), and reduced glutathione (GSH) levels were also measured. Myotubes morphology along with atrophic markers (calpain activity) were evaluated. Data show that in C2C12 myotubes, SB treatment reduced glucose consumption and uptake in a dose-dependent manner. SB inducing IR by disrupting insulin-mediated GLUT4 translocation in cultured myotubes. Alteration in oxidative stress-related markers (<em>i.e.</em> elevated ROS and LPO levels, reduced GSH) were also observed in SB-treated myotubes. Furthermore, decrease in the fusion index, length, and diameter of myotubes along with upregulation of calpain activity and decrease in muscle protein were also observed. Study concludes that SB exposure not only induced IR but also caused atrophy and oxidative stress in the C2C12 myotubes.</div></div>","PeriodicalId":251,"journal":{"name":"Biochimie","volume":"241 ","pages":"Pages 56-71"},"PeriodicalIF":3.0,"publicationDate":"2025-11-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145566653","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 : 2025-11-17DOI: 10.1016/j.biochi.2025.11.007
Lu-Fan Li, Yi-Man Liu, Yu-Jing Yan, Xing-Chen Wang, Le Chen, Jin-Hui Xu, Zhe Wang
Photoperiodic regulation exerts a significant influence on growth and development in seasonal mammals, yet its impact on oxidative stress and ferroptotic signaling in the Harderian glands of striped dwarf hamsters (Cricetulus barabensis) remains unresolved. Porphyrins, highly enriched in the Harderian glands of female rodents, undergo photodegradation under prolonged light exposure, generating reactive oxygen species (ROS) that may trigger ferroptosis. To elucidate photoperiod-dependent alterations, protoporphyrin levels, oxidative stress indicators, and ferroptosis-related markers were quantified in female hamsters maintained under long daylight (LD) and short daylight (SD) conditions. Under LD exposure, both δ-aminolevulinate synthase (ALA-S) and ferrochelatase (FECH) activities increased, while protoporphyrinogen oxidase (PPO) activity remained unchanged, collectively leading to elevated protoporphyrin accumulation. Concomitantly, LD hamsters exhibited increased hydrogen peroxide (H2O2) levels, enhanced superoxide dismutase (SOD) activity, and reduced systemic melatonin. LD exposure also elevated ferrous ion concentration while reducing ferritin protein expression. Notably, solute carrier family 7 member 11 (SLC7A11) expression increased under LD conditions, whereas acyl-CoA synthetase long-chain family member 4 (ACSL4) and glutathione peroxidase (GPx) activity remained stable. Malondialdehyde (MDA) levels did not differ between photoperiods. Collectively, these findings demonstrate that extended photoperiod promotes protoporphyrin accumulation and oxidative stress in the Harderian glands of female hamsters, but compensatory regulation of antioxidant enzymes and ferroptosis-related signaling proteins may attenuate lipid peroxidation and limit cellular injury.
{"title":"Long photoperiod induces porphyrin accumulation while maintaining lipid peroxidation stability in Harderian glands of female striped dwarf hamsters","authors":"Lu-Fan Li, Yi-Man Liu, Yu-Jing Yan, Xing-Chen Wang, Le Chen, Jin-Hui Xu, Zhe Wang","doi":"10.1016/j.biochi.2025.11.007","DOIUrl":"10.1016/j.biochi.2025.11.007","url":null,"abstract":"<div><div>Photoperiodic regulation exerts a significant influence on growth and development in seasonal mammals, yet its impact on oxidative stress and ferroptotic signaling in the Harderian glands of striped dwarf hamsters (<em>Cricetulus barabensis</em>) remains unresolved. Porphyrins, highly enriched in the Harderian glands of female rodents, undergo photodegradation under prolonged light exposure, generating reactive oxygen species (ROS) that may trigger ferroptosis. To elucidate photoperiod-dependent alterations, protoporphyrin levels, oxidative stress indicators, and ferroptosis-related markers were quantified in female hamsters maintained under long daylight (LD) and short daylight (SD) conditions. Under LD exposure, both δ-aminolevulinate synthase (ALA-S) and ferrochelatase (FECH) activities increased, while protoporphyrinogen oxidase (PPO) activity remained unchanged, collectively leading to elevated protoporphyrin accumulation. Concomitantly, LD hamsters exhibited increased hydrogen peroxide (H<sub>2</sub>O<sub>2</sub>) levels, enhanced superoxide dismutase (SOD) activity, and reduced systemic melatonin. LD exposure also elevated ferrous ion concentration while reducing ferritin protein expression. Notably, solute carrier family 7 member 11 (SLC7A11) expression increased under LD conditions, whereas acyl-CoA synthetase long-chain family member 4 (ACSL4) and glutathione peroxidase (GPx) activity remained stable. Malondialdehyde (MDA) levels did not differ between photoperiods. Collectively, these findings demonstrate that extended photoperiod promotes protoporphyrin accumulation and oxidative stress in the Harderian glands of female hamsters, but compensatory regulation of antioxidant enzymes and ferroptosis-related signaling proteins may attenuate lipid peroxidation and limit cellular injury.</div></div>","PeriodicalId":251,"journal":{"name":"Biochimie","volume":"241 ","pages":"Pages 98-105"},"PeriodicalIF":3.0,"publicationDate":"2025-11-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145558491","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 : 2025-11-17DOI: 10.1016/j.biochi.2025.11.008
Han Ba Bui , Duc Phu Phan, Wen-Guey Wu
Atragin and K-like are N-linked glycoproteins that belong to the P-III class of snake venom metalloproteinases, which hydrolyze components of the extracellular matrix and plasma proteins, thus contributing to vascular damage and coagulation disorders. In this study, we examined the effects of N-linked glycans on the proteolytic activity and structural stability of Atragin and K-like, which were purified from Naja atra cobra venom. Deglycosylation was performed using recombinant endo F3 under nondenaturing conditions. The removal of N-linked glycans significantly altered the proteolytic activity of both proteins against extracellular matrix substrates. In addition, neuraminidase-mediated desialylation markedly reduced the fibrinogenolytic activity of K-like. Free sialic acid binds to fibrinogen and inhibits its cleavage by K-like. Deglycosylation enhanced the heparin-binding affinity of Atragin and K-like. Heparin efficiently enhanced the proteolytic activity of both proteins irrespective of their glycosylation state. Deglycosylation reduced resistance to guanidine hydrochloride-induced unfolding, and K-like was more sensitive to thermal denaturation. These findings highlight the critical role of N-linked glycans in maintaining the enzymatic activity and structural integrity of venom metalloproteinases, and reveal how specific carbohydrate moieties, including terminal sialic acid residues, modulate their interactions with substrates. This study improves our understanding of venom proteins and may inform therapeutic strategies for snakebite treatment.
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