Pub Date : 2025-08-07DOI: 10.1016/j.ab.2025.115949
Jialong Tian , Pengli Lu , Huining Sha
Accurate identification of essential proteins is crucial in biomedical research. Traditional methods rely on Protein–Protein Interaction (PPI) networks and limited biological feature data, often neglecting the critical role of protein amino acid sequences. To address this, we propose the HCNS model, which integrates the Hypergraph Convolutional Network (HGCN) module, the Seq-CNN-MB-NAG feature extraction module, and the Multi-Layer Perceptron (MLP) recognition module, significantly enhancing the accuracy of essential protein identification. The HGCN module constructs a hypergraph from the PPI network and protein complex data, capturing the complex structural relationships between proteins. The Seq-CNN-MB-NAG module utilizes Convolutional Neural Networks (CNN), Multi-Head Self-Attention (MHSA), Bidirectional Long Short-Term Memory (Bi-LSTM), and NAG Transformer to process protein amino acid sequences and extract sequence features. The MLP module then fuses these two sets of features for precise recognition. Experimental results show that the HCNS model outperforms existing methods, achieving an accuracy of 93.38%, with an Area Under the Curve (AUC) of 98.33% and an Area Under the Precision–Recall Curve (AUPR) of 97.16%, demonstrating its potential in essential protein identification.
准确鉴定必需蛋白质在生物医学研究中至关重要。传统的方法依赖于蛋白质-蛋白质相互作用（PPI）网络和有限的生物学特征数据，往往忽略了蛋白质氨基酸序列的关键作用。为了解决这个问题，我们提出了HCNS模型，该模型集成了超图卷积网络（HGCN）模块、Seq-CNN-MB-NAG特征提取模块和多层感知器（MLP）识别模块，显著提高了必需蛋白质识别的准确性。HGCN模块利用PPI网络和蛋白质复合体数据构建超图，捕捉蛋白质之间复杂的结构关系。Seq-CNN-MB-NAG模块利用卷积神经网络（CNN）、多头自注意（MHSA）、双向长短期记忆（Bi-LSTM）和NAG Transformer处理蛋白质氨基酸序列并提取序列特征。然后，MLP模块融合这两组特征以进行精确识别。实验结果表明，HCNS模型的准确率为93.38%，曲线下面积（Area Under the Curve， AUC）为98.33%，精确召回曲线下面积（Area Under Precision-Recall Curve， AUPR）为97.16%，优于现有的方法，显示了其在必需蛋白鉴定中的潜力。
{"title":"HCNS:A deep learning model for identifying essential proteins based on hypergraph convolution and sequence features","authors":"Jialong Tian , Pengli Lu , Huining Sha","doi":"10.1016/j.ab.2025.115949","DOIUrl":"10.1016/j.ab.2025.115949","url":null,"abstract":"<div><div>Accurate identification of essential proteins is crucial in biomedical research. Traditional methods rely on Protein–Protein Interaction (PPI) networks and limited biological feature data, often neglecting the critical role of protein amino acid sequences. To address this, we propose the HCNS model, which integrates the Hypergraph Convolutional Network (HGCN) module, the Seq-CNN-MB-NAG feature extraction module, and the Multi-Layer Perceptron (MLP) recognition module, significantly enhancing the accuracy of essential protein identification. The HGCN module constructs a hypergraph from the PPI network and protein complex data, capturing the complex structural relationships between proteins. The Seq-CNN-MB-NAG module utilizes Convolutional Neural Networks (CNN), Multi-Head Self-Attention (MHSA), Bidirectional Long Short-Term Memory (Bi-LSTM), and NAG Transformer to process protein amino acid sequences and extract sequence features. The MLP module then fuses these two sets of features for precise recognition. Experimental results show that the HCNS model outperforms existing methods, achieving an accuracy of 93.38%, with an Area Under the Curve (AUC) of 98.33% and an Area Under the Precision–Recall Curve (AUPR) of 97.16%, demonstrating its potential in essential protein identification.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"707 ","pages":"Article 115949"},"PeriodicalIF":2.5,"publicationDate":"2025-08-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144809717","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-08-06DOI: 10.1016/j.ab.2025.115952
Tasmin Karim , Md Shazzad Hossain Shaon , Md Mamun Ali , Sobhy M. Ibrahim , Mst Shapna Akter , Kawsar Ahmed , Francis M. Bui , Li Chen , Mohammad Ali Moni
Protein S-nitrosylation (SNO) is a process involving the covalent modification of cysteine residues by nitric oxide (NO) and its derivatives. Numerous studies have demonstrated that SNO is significantly involved in cell function and pathophysiology. The identification of SNO sites is significant in clarifying their function in cellular physiology, disease processes, and potential treatment strategies, rendering it of paramount importance in medical science. This study developed a machine learning (ML) model named “K-SNOpred” and found notable performance in identifying SNO sites using the Latent Semantic Analysis (LSA) feature embedding system. After collecting dbSNO and RecSNO datasets from the literature search, we applied three feature embedding systems: Doc2vec, FastText, and LSA on each dataset. The study employed various ML models and assessed their performance using multiple evaluation metrics through independent testing and 10-fold cross-validation. The evaluation's outcomes demonstrate that the proposed model achieved an accuracy of 87.56 % and an AUC score of 95.06 %, outperforming existing state-of-the-art (SOTA) models by nearly 10 % in accuracy and 6 % in AUC. Furthermore, the model demonstrated balanced sensitivity and specificity, indicating its ability to detect both positive and negative SNO sites accurately. The outstanding performance of the K-SNOpred model demonstrates its high potential for clinical use and its applicability in the biotechnology field.
{"title":"K-SNOpred: Identification of protein S-nitrosylation sites through word embedding features and machine learning","authors":"Tasmin Karim , Md Shazzad Hossain Shaon , Md Mamun Ali , Sobhy M. Ibrahim , Mst Shapna Akter , Kawsar Ahmed , Francis M. Bui , Li Chen , Mohammad Ali Moni","doi":"10.1016/j.ab.2025.115952","DOIUrl":"10.1016/j.ab.2025.115952","url":null,"abstract":"<div><div>Protein S-nitrosylation (SNO) is a process involving the covalent modification of cysteine residues by nitric oxide (NO) and its derivatives. Numerous studies have demonstrated that SNO is significantly involved in cell function and pathophysiology. The identification of SNO sites is significant in clarifying their function in cellular physiology, disease processes, and potential treatment strategies, rendering it of paramount importance in medical science. This study developed a machine learning (ML) model named “K-SNOpred” and found notable performance in identifying SNO sites using the Latent Semantic Analysis (LSA) feature embedding system. After collecting dbSNO and RecSNO datasets from the literature search, we applied three feature embedding systems: Doc2vec, FastText, and LSA on each dataset. The study employed various ML models and assessed their performance using multiple evaluation metrics through independent testing and 10-fold cross-validation. The evaluation's outcomes demonstrate that the proposed model achieved an accuracy of 87.56 % and an AUC score of 95.06 %, outperforming existing state-of-the-art (SOTA) models by nearly 10 % in accuracy and 6 % in AUC. Furthermore, the model demonstrated balanced sensitivity and specificity, indicating its ability to detect both positive and negative SNO sites accurately. The outstanding performance of the K-SNOpred model demonstrates its high potential for clinical use and its applicability in the biotechnology field.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"707 ","pages":"Article 115952"},"PeriodicalIF":2.5,"publicationDate":"2025-08-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144803266","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
In this study, olfactory receptors (ORs) and Golf proteins fused with split luciferase were examined to develop a sensitive assay for identifying OR ligands. Although specific luminescence was observed from HEK293T cells expressing both fusion proteins (OR1A1-LgBiT and SmBiT-Mini-Golf) in response to an OR1A1 ligand, its intensity was low. To improve it, a fusion protein (V1N-G4S3-SmBiT-G4S3-Mini-Golf), was created where V1N contained the palmitoylation site necessary for localizing Mini-Golf to the cell membrane. When OR1A1-LgBiT and V1N-G4S3-SmBiT-G4S3-Mini-Golf were co-expressed in HEK293T cells, specific luminescence by ligand stimulation was considerably increased. The cell surface expression of OR1A1-LgBiT was markedly lower than that of OR1A1. Co-expression of receptor transporting protein 1 short form (RTP1S) promoted the cell surface expression of OR1A1-LgBiT more efficiently than OR1A1. Prediction using AlphaFold3 suggested that the N-terminal domain of RTP1S interacts with LgBiT in OR1A1-LgBiT. Co-expression of OR1A1-LgBiT, V1N-G4S3-SmBiT-G4S3-Mini-Golf, and RTP1S in HEK293T cells enhanced the sensitivity for detecting at least some ligands by 1000- to 3000-fold compared to the conventional cAMP assay. These findings suggest that optimizing the assembly of OR1A1-LgBiT and SmBiT-Mini-Golf at the cell membrane is essential for developing a sensitive assay. The OR response-detection system employing split luciferase holds significant potential for investigating ligands of orphan ORs.
{"title":"A sensitive split luciferase assay for detecting olfactory receptor response to odorants","authors":"Natsumi Tanazawa , Sho Obayashi , Shuji Hinuma , Shun'ichi Kuroda","doi":"10.1016/j.ab.2025.115954","DOIUrl":"10.1016/j.ab.2025.115954","url":null,"abstract":"<div><div>In this study, olfactory receptors (ORs) and Golf proteins fused with split luciferase were examined to develop a sensitive assay for identifying OR ligands. Although specific luminescence was observed from HEK293T cells expressing both fusion proteins (OR1A1-LgBiT and SmBiT-Mini-Golf) in response to an OR1A1 ligand, its intensity was low. To improve it, a fusion protein (V1N-G4S3-SmBiT-G4S3-Mini-Golf), was created where V1N contained the palmitoylation site necessary for localizing Mini-Golf to the cell membrane. When OR1A1-LgBiT and V1N-G4S3-SmBiT-G4S3-Mini-Golf were co-expressed in HEK293T cells, specific luminescence by ligand stimulation was considerably increased. The cell surface expression of OR1A1-LgBiT was markedly lower than that of OR1A1. Co-expression of receptor transporting protein 1 short form (RTP1S) promoted the cell surface expression of OR1A1-LgBiT more efficiently than OR1A1. Prediction using AlphaFold3 suggested that the N-terminal domain of RTP1S interacts with LgBiT in OR1A1-LgBiT. Co-expression of OR1A1-LgBiT, V1N-G4S3-SmBiT-G4S3-Mini-Golf, and RTP1S in HEK293T cells enhanced the sensitivity for detecting at least some ligands by 1000- to 3000-fold compared to the conventional cAMP assay. These findings suggest that optimizing the assembly of OR1A1-LgBiT and SmBiT-Mini-Golf at the cell membrane is essential for developing a sensitive assay. The OR response-detection system employing split luciferase holds significant potential for investigating ligands of orphan ORs.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"707 ","pages":"Article 115954"},"PeriodicalIF":2.5,"publicationDate":"2025-08-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144798011","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-08-04DOI: 10.1016/j.ab.2025.115953
Jyoti Jaiswal, Ashish P. Singh, Rajeshwar P. Sinha
C-phycocyanin is a natural blue-colored pigment-protein complex existing as phycobiliprotein in cyanobacteria. C-phycocyanin is made up of two different subunits of α- and -β monomers. These subunits assemble to form a cylindrical structure known as phycocyanobilin chromophore. In the present study, C-phycocyanin was purified from the cyanobacterium Spirulina subsalsa HKAR-19 using ammonium sulfate precipitation followed by sucrose density gradient ultracentrifugation methods and characterized by various spectroscopic techniques. The purified C-phycocyanin showed an absorption peak at 615 nm. A characteristic fluorescence emission peak at 642 nm when excited over 615 nm, indicates the integrity and functionality of protein structure. The scavenging activity against free radicals of DPPH, ABTS, and superoxide (O2·−) was evaluated by using free radical scavenging assays. C-phycocyanin showed dose-dependent in vitro antioxidative properties towards free radicals. It possesses higher antioxidant activity toward ABTS•+ and DPPH•. Due to its high antioxidant activity, it can be used as a colorant and therapeutic agent against oxidative stress.
c -藻蓝蛋白是一种天然的蓝色色素蛋白复合物，以藻胆蛋白的形式存在于蓝藻细菌中。c -藻蓝蛋白由α-和-β单体的两个不同亚基组成。这些亚基组合形成一个圆柱形结构，称为藻蓝胆素发色团。本研究采用硫酸铵沉淀-蔗糖密度梯度超离心的方法从藻蓝杆菌下螺旋藻HKAR-19中纯化出c -藻蓝蛋白，并用各种光谱技术对其进行了表征。纯化后的c -藻蓝蛋白在615 nm处有吸收峰。在615 nm以上激发时，在642 nm处有一个特征荧光发射峰，表明蛋白质结构的完整性和功能性。通过自由基清除试验评估其对DPPH、ABTS和超氧化物（O2·−）自由基的清除活性。c -藻蓝蛋白对自由基的体外抗氧化性能表现出剂量依赖性。对ABTS•+和DPPH•具有较高的抗氧化活性。由于其高抗氧化活性，它可以用作着色剂和抗氧化应激治疗剂。
{"title":"Characterization and in vitro antioxidant activity of C-phycocyanin from the cyanobacterium Spirulina subsalsa HKAR-19","authors":"Jyoti Jaiswal, Ashish P. Singh, Rajeshwar P. Sinha","doi":"10.1016/j.ab.2025.115953","DOIUrl":"10.1016/j.ab.2025.115953","url":null,"abstract":"<div><div>C-phycocyanin is a natural blue-colored pigment-protein complex existing as phycobiliprotein in cyanobacteria. C-phycocyanin is made up of two different subunits of α- and -β monomers. These subunits assemble to form a cylindrical structure known as phycocyanobilin chromophore. In the present study, C-phycocyanin was purified from the cyanobacterium <em>Spirulina subsalsa</em> HKAR-19 using ammonium sulfate precipitation followed by sucrose density gradient ultracentrifugation methods and characterized by various spectroscopic techniques. The purified C-phycocyanin showed an absorption peak at 615 nm. A characteristic fluorescence emission peak at 642 nm when excited over 615 nm, indicates the integrity and functionality of protein structure. The scavenging activity against free radicals of DPPH, ABTS, and superoxide (O<sub>2</sub>·<sup>−</sup>) was evaluated by using free radical scavenging assays. C-phycocyanin showed dose-dependent <em>in vitro</em> antioxidative properties towards free radicals. It possesses higher antioxidant activity toward <em>ABTS</em><sup>•+</sup> and <em>DPPH</em><sup>•</sup>. Due to its high antioxidant activity, it can be used as a colorant and therapeutic agent against oxidative stress.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"706 ","pages":"Article 115953"},"PeriodicalIF":2.5,"publicationDate":"2025-08-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144766888","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Gastric cancer (GC) is one of the leading causes of cancer-related mortality worldwide, with late-stage diagnosis and limited treatment efficacy. The major challenges in chemotherapy and radiotherapy for GC include drug resistance, systemic toxicity, limited tumor selectivity, adverse side effects, tumor heterogeneity, and the development of radioresistance, all of which hinder treatment efficacy and patient outcomes. Aptamers, single-stranded DNA or RNA molecules with high specificity and affinity for molecular targets, offer a promising alternative to traditional diagnostic and therapeutic approaches. Due to their high specificity and low immunogenicity, aptamers improve non-invasive diagnostic techniques, enabling early GC detection and targeted therapeutic applications. Therapeutically, aptamers facilitate precise drug targeting, reducing systemic toxicity and improving treatment outcomes, especially for drug-resistant and metastatic cases. Integration with nanotechnology further enhances their potential, enabling theranostic applications that combine diagnosis and therapy. Despite these advancements, challenges such as physiological stability, renal clearance, and tissue permeability remain, necessitating further research into chemical modifications and nanocarrier-based delivery systems. This study provides a comprehensive review of recent advancements in aptamer technology and its evolving applications in GC, with a particular emphasis on diagnostic and therapeutic strategies. The first section explores aptamer development strategies, including emerging selection techniques and molecular modifications to enhance their specificity and stability. The subsequent section examines the role of aptamers in GC diagnostics and treatment, with a focus on targeted chemotherapy and immunotherapy. Lastly, we discuss the current challenges in aptamer-based approaches and outline future perspectives for their clinical translation.
{"title":"Advances in aptamer technology and its updated applications in gastric cancer","authors":"Sayedeh Azimeh Hosseini , Arghavan Rakhshani Nejad , Hassan Valadbeigi , Mohammad Hossein Haddadi , Monireh Bazdar","doi":"10.1016/j.ab.2025.115950","DOIUrl":"10.1016/j.ab.2025.115950","url":null,"abstract":"<div><div>Gastric cancer (GC) is one of the leading causes of cancer-related mortality worldwide, with late-stage diagnosis and limited treatment efficacy. The major challenges in chemotherapy and radiotherapy for GC include drug resistance, systemic toxicity, limited tumor selectivity, adverse side effects, tumor heterogeneity, and the development of radioresistance, all of which hinder treatment efficacy and patient outcomes. Aptamers, single-stranded DNA or RNA molecules with high specificity and affinity for molecular targets, offer a promising alternative to traditional diagnostic and therapeutic approaches. Due to their high specificity and low immunogenicity, aptamers improve non-invasive diagnostic techniques, enabling early GC detection and targeted therapeutic applications. Therapeutically, aptamers facilitate precise drug targeting, reducing systemic toxicity and improving treatment outcomes, especially for drug-resistant and metastatic cases. Integration with nanotechnology further enhances their potential, enabling theranostic applications that combine diagnosis and therapy. Despite these advancements, challenges such as physiological stability, renal clearance, and tissue permeability remain, necessitating further research into chemical modifications and nanocarrier-based delivery systems. This study provides a comprehensive review of recent advancements in aptamer technology and its evolving applications in GC, with a particular emphasis on diagnostic and therapeutic strategies. The first section explores aptamer development strategies, including emerging selection techniques and molecular modifications to enhance their specificity and stability. The subsequent section examines the role of aptamers in GC diagnostics and treatment, with a focus on targeted chemotherapy and immunotherapy. Lastly, we discuss the current challenges in aptamer-based approaches and outline future perspectives for their clinical translation.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"706 ","pages":"Article 115950"},"PeriodicalIF":2.5,"publicationDate":"2025-08-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144764029","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-07-30DOI: 10.1016/j.ab.2025.115951
Xia Li , Qi Chen , Yating Zhu , Haofei Hong , Zhifang Zhou , Zhimeng Wu , Jie Shi
Rapid and accurate detection of neutralizing antibody (nAb) is essential for assessing vaccine efficacy and immune protection against infectious diseases. In the present study, we developed a neutralization-antibody detection qPCR platform (NAD-qPCR) that quantifies nAb potency by integrating antigen-receptor binding specificity with qPCR sensitivity. Using the SARS-CoV-2 nAb as a proof-of-concept target, this system comprised a hybrid probe with the viral RBD as an antigen module covalently conjugated to a reporter DNA, and magnetic beads functionalized with the ACE2 mimic mini-protein as a receptor module, thereby recapitulating the natural antigen-receptor interactions. With this design, the neutralizing capability of nAb against the antigen was transformed into a competitive reduction of the probe binding and DNA amplification signal by qPCR. The NAD-qPCR demonstrated robust performance in quantifying commercial SARS-CoV-2 nAb with a LOD of 4 ng/μL. Furthermore, it was demonstrated to be capable of effectively profiling a large number of vaccinated donor serum with broad neutralization activity. Notably, the hybrid probe and the capture beads were designed modularly, which allows for straightforward adaptation to other pathogens by replacing the module component. This platform offers a rapid, high-throughput, and universally applicable approach for nAb detection in vaccine development and immune monitoring.
{"title":"A modular and universal platform for neutralizing-antibody profiling by quantifying antigen-receptor interactions via qPCR","authors":"Xia Li , Qi Chen , Yating Zhu , Haofei Hong , Zhifang Zhou , Zhimeng Wu , Jie Shi","doi":"10.1016/j.ab.2025.115951","DOIUrl":"10.1016/j.ab.2025.115951","url":null,"abstract":"<div><div>Rapid and accurate detection of neutralizing antibody (nAb) is essential for assessing vaccine efficacy and immune protection against infectious diseases. In the present study, we developed a neutralization-antibody detection qPCR platform (NAD-qPCR) that quantifies nAb potency by integrating antigen-receptor binding specificity with qPCR sensitivity. Using the SARS-CoV-2 nAb as a proof-of-concept target, this system comprised a hybrid probe with the viral RBD as an antigen module covalently conjugated to a reporter DNA, and magnetic beads functionalized with the ACE2 mimic mini-protein as a receptor module, thereby recapitulating the natural antigen-receptor interactions. With this design, the neutralizing capability of nAb against the antigen was transformed into a competitive reduction of the probe binding and DNA amplification signal by qPCR. The NAD-qPCR demonstrated robust performance in quantifying commercial SARS-CoV-2 nAb with a LOD of 4 ng/μL. Furthermore, it was demonstrated to be capable of effectively profiling a large number of vaccinated donor serum with broad neutralization activity. Notably, the hybrid probe and the capture beads were designed modularly, which allows for straightforward adaptation to other pathogens by replacing the module component. This platform offers a rapid, high-throughput, and universally applicable approach for nAb detection in vaccine development and immune monitoring.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"706 ","pages":"Article 115951"},"PeriodicalIF":2.5,"publicationDate":"2025-07-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144756934","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-07-18DOI: 10.1016/j.ab.2025.115947
Ulrike Vogel, Renger Dijkstra, Koen Beerens, Tom Desmet
CDP-tyvelose 2-epimerase-like enzymes and UDP-glucose 4-epimerases belong to the Nucleotide Sugar active Short-Chain Dehydrogenase/Reductase (NS-SDR) superfamily and convert NDP-Glc to NDP-Man and NDP-Gal, respectively. The product detection is well established using HPLC-based methods, capillary electrophoresis and thin layer chromatography for both epimerization reactions. However, the lack of a reliable colorimetric assays and poor substrate availability slow down the screening process of these enzymes, for example during enzyme engineering projects. After testing 16 phenolic compounds with different aldohexoses in concentrated sulfuric acid, thymol was found suitable for the detection of (NDP-) mannose and 1-naphthol for the detection of (NDP-) galactose in the presence of (NDP-) glucose. As proof-of-concept, the optimized assay was used for the biochemical characterization of the CDP-tyvelose 2-epimerase from Ardenticatenia bacterium.
{"title":"Colorimetric assays for the detection of C4- and C2-epimerization of NDP-glucose to NDP-galactose and NDP-mannose","authors":"Ulrike Vogel, Renger Dijkstra, Koen Beerens, Tom Desmet","doi":"10.1016/j.ab.2025.115947","DOIUrl":"10.1016/j.ab.2025.115947","url":null,"abstract":"<div><div>CDP-tyvelose 2-epimerase-like enzymes and UDP-glucose 4-epimerases belong to the Nucleotide Sugar active Short-Chain Dehydrogenase/Reductase (NS-SDR) superfamily and convert NDP-Glc to NDP-Man and NDP-Gal, respectively. The product detection is well established using HPLC-based methods, capillary electrophoresis and thin layer chromatography for both epimerization reactions. However, the lack of a reliable colorimetric assays and poor substrate availability slow down the screening process of these enzymes, for example during enzyme engineering projects. After testing 16 phenolic compounds with different aldohexoses in concentrated sulfuric acid, thymol was found suitable for the detection of (NDP-) mannose and 1-naphthol for the detection of (NDP-) galactose in the presence of (NDP-) glucose. As proof-of-concept, the optimized assay was used for the biochemical characterization of the CDP-tyvelose 2-epimerase from <em>Ardenticatenia bacterium</em>.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"706 ","pages":"Article 115947"},"PeriodicalIF":2.6,"publicationDate":"2025-07-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144671018","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Previous Mendelian randomization (MR) studies have explored the role of plasma proteins in ulcerative colitis (UC), but the underlying mechanisms, particularly how plasma proteins influence UC risk via immune-mediated pathways, remain unclear. This study integrates two-sample MR and mediation analysis to investigate whether plasma proteins influence UC risk through antibody-mediated immune responses, aiming to identify novel biomarkers and therapeutic targets.
Methods
We analyzed 4907 plasma proteins from a genome-wide association study, 46 immune antibody responses, and UC data from the FinnGen consortium. Two-sample MR was used to assess causal relationships among plasma proteins, antibody responses, and UC. Mediation MR analysis was conducted to evaluate whether specific antibody responses mediate the effect of plasma proteins on UC risk.
Results
A total of 80 plasma proteins with significant causal associations with UC were identified (P < 0.05). Two antibody responses, Epstein-Barr virus (EBV) EA-D antibody levels and anti-HSV-1 IgG seropositivity, were significantly and inversely associated with UC risk (EBV: OR = 0.794, 95 % CI: 0.646–0.974, P = 0.027; HSV-1 IgG: OR = 0.891, 95 % CI: 0.801–0.992, P = 0.035). Five plasma proteins showed significant causal effects on these antibody responses: UBC, TIMD4, and NEFL (linked to EBV EA-D), and TMEM70 and HIF1A (linked to HSV-1 IgG). Mediation analysis revealed that antibody responses explained 9.2 %–13.2 % of the total effects of these proteins on UC risk. For example, UBC increased UC risk partially through reduced EBV EA-D antibodies (mediated effect = 0.0588), while HIF1A contributed to UC risk via suppressed HSV-1 IgG seropositivity.
Conclusion
This study identifies a novel immuno-genetic pathway in UC pathogenesis, where specific plasma proteins influence disease risk through modulation of viral antibody responses. These findings suggest potential targets, such as UBC, HIF1A, and TIMD4, for biomarker development and immune-focused interventions in UC.
先前的孟德尔随机化（MR）研究已经探索了血浆蛋白在溃疡性结肠炎（UC）中的作用，但其潜在机制，特别是血浆蛋白如何通过免疫介导途径影响UC风险尚不清楚。本研究结合两样本MR和中介分析，探讨血浆蛋白是否通过抗体介导的免疫反应影响UC风险，旨在发现新的生物标志物和治疗靶点。方法我们分析了来自全基因组关联研究的4907种血浆蛋白、46种免疫抗体应答和来自FinnGen联盟的UC数据。双样本MR用于评估血浆蛋白、抗体反应和UC之间的因果关系。进行中介MR分析以评估特异性抗体反应是否介导血浆蛋白对UC风险的影响。结果共鉴定出80种与UC有显著因果关系的血浆蛋白(P <；0.05)。eb病毒（EBV） EA-D抗体水平和抗hsv -1 IgG血清阳性与UC风险呈显著负相关(EBV: OR = 0.794, 95% CI: 0.646-0.974, P = 0.027；1型单纯疱疹病毒免疫球蛋白:= 0.891,95%置信区间CI: 0.801 - -0.992, P = 0.035)。五种血浆蛋白对这些抗体反应有显著的因果影响：UBC、TIMD4和NEFL（与EBV EA-D相关），以及TMEM70和HIF1A（与HSV-1 IgG相关）。中介分析显示，抗体反应解释了这些蛋白质对UC风险的总影响的9.2% - 13.2%。例如，UBC部分通过降低EBV EA-D抗体增加UC风险（介导效应= 0.0588），而HIF1A通过抑制HSV-1 IgG血清阳性增加UC风险。结论本研究确定了UC发病机制中的一种新的免疫遗传途径，其中特异性血浆蛋白通过调节病毒抗体反应影响疾病风险。这些发现提示了UC生物标志物开发和免疫干预的潜在靶点，如UBC、HIF1A和TIMD4。
{"title":"Decoding ulcerative colitis: Plasma protein-mediated antibody immune responses influence disease risk","authors":"Xuyong Chen, Haidong Wu, Liudan Wang, Yifan Guo, Xinpu Miao","doi":"10.1016/j.ab.2025.115946","DOIUrl":"10.1016/j.ab.2025.115946","url":null,"abstract":"<div><h3>Objective</h3><div>Previous Mendelian randomization (MR) studies have explored the role of plasma proteins in ulcerative colitis (UC), but the underlying mechanisms, particularly how plasma proteins influence UC risk via immune-mediated pathways, remain unclear. This study integrates two-sample MR and mediation analysis to investigate whether plasma proteins influence UC risk through antibody-mediated immune responses, aiming to identify novel biomarkers and therapeutic targets.</div></div><div><h3>Methods</h3><div>We analyzed 4907 plasma proteins from a genome-wide association study, 46 immune antibody responses, and UC data from the FinnGen consortium. Two-sample MR was used to assess causal relationships among plasma proteins, antibody responses, and UC. Mediation MR analysis was conducted to evaluate whether specific antibody responses mediate the effect of plasma proteins on UC risk.</div></div><div><h3>Results</h3><div>A total of 80 plasma proteins with significant causal associations with UC were identified (P < 0.05). Two antibody responses, Epstein-Barr virus (EBV) EA-D antibody levels and anti-HSV-1 IgG seropositivity, were significantly and inversely associated with UC risk (EBV: OR = 0.794, 95 % CI: 0.646–0.974, P = 0.027; HSV-1 IgG: OR = 0.891, 95 % CI: 0.801–0.992, P = 0.035). Five plasma proteins showed significant causal effects on these antibody responses: UBC, TIMD4, and NEFL (linked to EBV EA-D), and TMEM70 and HIF1A (linked to HSV-1 IgG). Mediation analysis revealed that antibody responses explained 9.2 %–13.2 % of the total effects of these proteins on UC risk. For example, UBC increased UC risk partially through reduced EBV EA-D antibodies (mediated effect = 0.0588), while HIF1A contributed to UC risk via suppressed HSV-1 IgG seropositivity.</div></div><div><h3>Conclusion</h3><div>This study identifies a novel immuno-genetic pathway in UC pathogenesis, where specific plasma proteins influence disease risk through modulation of viral antibody responses. These findings suggest potential targets, such as UBC, HIF1A, and TIMD4, for biomarker development and immune-focused interventions in UC.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"706 ","pages":"Article 115946"},"PeriodicalIF":2.6,"publicationDate":"2025-07-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144654331","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-07-12DOI: 10.1016/j.ab.2025.115943
Alexander Spreinat , Carola Wilczek , Christin Ronsör , Andrea Ernst
Lateral Flow Assays (LFA) have been valuable tools for point-of-care diagnostics for decades. During periods of high testing demand, as recently in the Covid-19-pandemic, it is necessary to deepen the understanding of test and control line signal intensities. This study investigates the interaction between proteins and structurally different nitrocellulose (CN) membranes and the resulting effects on signal intensity in an LFA under the influence of various buffer additives. The experiments focused on quantitative protein adsorption, protein stability, protein line printing and changes in signal intensity in a human chorionic gonadotropin (hCG)-assay. A method was established for detecting the widths and intensities of fluorescent protein lines. We were able to show that changes in signal intensity of LFAs are driven by accessibility of the antibodies, by hydrophilicity of the membrane or assisted adsorption of antibodies onto the membrane. Additionally, the inclusion of sodium chloride, polysorbate 80 and sodium dodecylbenzenesulfonate can enhance signal intensity. The method developed for protein line analysis has proven to be effective and can help to understand protein-membrane-interactions on a macroscopic level. We demonstrate that LFA-manufacturers have a range of options to fine-tune assay performance without major modifications to assay components.
{"title":"Enhancing lateral flow assay performance: Buffer additives and protein-membrane interactions","authors":"Alexander Spreinat , Carola Wilczek , Christin Ronsör , Andrea Ernst","doi":"10.1016/j.ab.2025.115943","DOIUrl":"10.1016/j.ab.2025.115943","url":null,"abstract":"<div><div>Lateral Flow Assays (LFA) have been valuable tools for point-of-care diagnostics for decades. During periods of high testing demand, as recently in the Covid-19-pandemic, it is necessary to deepen the understanding of test and control line signal intensities. This study investigates the interaction between proteins and structurally different nitrocellulose (CN) membranes and the resulting effects on signal intensity in an LFA under the influence of various buffer additives. The experiments focused on quantitative protein adsorption, protein stability, protein line printing and changes in signal intensity in a human chorionic gonadotropin (hCG)-assay. A method was established for detecting the widths and intensities of fluorescent protein lines. We were able to show that changes in signal intensity of LFAs are driven by accessibility of the antibodies, by hydrophilicity of the membrane or assisted adsorption of antibodies onto the membrane. Additionally, the inclusion of sodium chloride, polysorbate 80 and sodium dodecylbenzenesulfonate can enhance signal intensity. The method developed for protein line analysis has proven to be effective and can help to understand protein-membrane-interactions on a macroscopic level. We demonstrate that LFA-manufacturers have a range of options to fine-tune assay performance without major modifications to assay components.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"706 ","pages":"Article 115943"},"PeriodicalIF":2.6,"publicationDate":"2025-07-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144631456","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pub Date : 2025-07-11DOI: 10.1016/j.ab.2025.115945
Felice C. Simeone
Hallachrome is a 1,2-anthraquinone secreted by marine polychaete worms whose toxicity and antimicrobial properties have been reported; the origin of this biological activity, however, remains elusive. Voltammetric studies reveal reversible redox behavior in the pH range 1–10, consistent with a two-electron, two-proton (2e−/2H+) mechanism. The electron-donating substituents in hallachrome (hydroxyl and methyl groups) stabilize the protonated hydroquinone form and result in a substantial cathodic shift of 0.4–0.5 V compared to unsubstituted 1,2-benzoquinone.
The electrochemical analysis reveals that hallachrome (redox potential −0.12 V vs. SHE at pH 7) might be capable, from a thermodynamic point of view, of oxidizing key cellular antioxidants, including NADH (−0.32 V), NADPH (−0.32 V), and glutathione (−0.24 V). These findings provide fundamental insights into the structure-activity relationships governing quinone electrochemistry and establish a foundation for understanding hallachrome's biological activity.
Hallachrome是一种由海洋多毛类蠕虫分泌的1,2-蒽醌，其毒性和抗菌特性已被报道；然而，这种生物活性的起源仍然难以捉摸。伏安研究表明，在pH值1 ~ 10范围内，其氧化还原行为是可逆的，符合双电子、双质子（2e-/2H+）机制。hallachrome中的给电子取代基（羟基和甲基）稳定了质子化对苯二酚的形式，与未取代的1,2-苯醌相比，导致了0.4-0.5 V的阴极位移。电化学分析表明，从热力学角度来看，hallachrome （pH = 7时氧化还原电位-0.12 V vs. SHE）可能能够氧化关键的细胞抗氧化剂，包括NADH (-0.32 V), NADPH （-0.32 V）和谷胱甘肽（-0.24 V）。这些发现为研究醌类电化学的构效关系提供了基础，并为进一步了解hallachrome的生物活性奠定了基础。
{"title":"Proton-coupled electron transfer in hallachrome, a natural 1,2 anthraquinone: Linking electrochemical properties to biological activity","authors":"Felice C. Simeone","doi":"10.1016/j.ab.2025.115945","DOIUrl":"10.1016/j.ab.2025.115945","url":null,"abstract":"<div><div>Hallachrome is a 1,2-anthraquinone secreted by marine polychaete worms whose toxicity and antimicrobial properties have been reported; the origin of this biological activity, however, remains elusive. Voltammetric studies reveal reversible redox behavior in the pH range 1–10, consistent with a two-electron, two-proton (2e<sup>−</sup>/2H<sup>+</sup>) mechanism. The electron-donating substituents in hallachrome (hydroxyl and methyl groups) stabilize the protonated hydroquinone form and result in a substantial cathodic shift of 0.4–0.5 V compared to unsubstituted 1,2-benzoquinone.</div><div>The electrochemical analysis reveals that hallachrome (redox potential −0.12 V vs. SHE at pH 7) might be capable, from a thermodynamic point of view, of oxidizing key cellular antioxidants, including NADH (−0.32 V), NADPH (−0.32 V), and glutathione (−0.24 V). These findings provide fundamental insights into the structure-activity relationships governing quinone electrochemistry and establish a foundation for understanding hallachrome's biological activity.</div></div>","PeriodicalId":7830,"journal":{"name":"Analytical biochemistry","volume":"706 ","pages":"Article 115945"},"PeriodicalIF":2.6,"publicationDate":"2025-07-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144625290","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
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