Biochimica et biophysica acta. Proteins and proteomics最新文献_第2页

Flexibility in acidophilic thioredoxins: Insights from Asp43 substitutions in E. coli thioredoxin 嗜酸硫氧还蛋白的灵活性：来自大肠杆菌硫氧还蛋白Asp43取代的见解。

IF 2.3 4区生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et biophysica acta. Proteins and proteomics

Pub Date : 2025-11-19 DOI: 10.1016/j.bbapap.2025.141115

Oanh Mai Ho, Mohammed Shazaly A. Elhassan, Khang Nguyen, ChangWoo Lee

Thioredoxins (Trxs) are small, highly conserved oxidoreductases characterized by a redox-active CXXC motif. While the structural adaptation of modern Trxs such as Escherichia coli Trx (EcTrx) has been described, the mechanisms underlying Trx adaptation to acidic environments remain unclear. In EcTrx, Asp43 stabilizes the active-site region through a water-mediated hydrogen bond with Lys57, which modulates the protonation state of Cys32 in cooperation with Asp26. Comparative sequence analysis shows that small, nonpolar amino acids—such as Gly and Ala—are frequently found at position 43 in acidophilic Trxs, indicating that structural flexibility at this position may contribute to acidic adaptation. To test how substitutions at position 43 affect Trx stability and function, we generated EcTrx mutants (D43G, D43A, D43S, D43N, D43L, and D43E). Thermal shift and guanidinium chloride-induced unfolding assays revealed that D43A and D43S markedly reduced stability, while D43G retained moderate stability, likely due to tighter N-terminal packing as observed in the acidophilic Acetobacter aceti Trx. These three mutants also displayed increased conformational flexibility, whereas D43N and D43L conferred partial stabilization through polar or hydrophobic side chains. In contrast, D43E—mimicking ancestral Glu—restored hydrogen bonding and enhanced thermal stability, resulting in the most rigid structure. Most mutants retained catalytic activity in DTNB assays, except D43S, which showed 50% of wild-type activity. Overall, our results demonstrate that Asp43 is critical for maintaining EcTrx structural stability and suggest that enhanced, but not excessive, flexibility at this position facilitates Trx adaptation to acidic environments.

硫氧还毒素（Trxs）是一种小而高度保守的氧化还原酶，其特征是具有氧化还原活性的CXXC基序。虽然现代Trx如大肠杆菌Trx （EcTrx）的结构适应性已经被描述，但Trx适应酸性环境的机制尚不清楚。在EcTrx中，Asp43通过水介导的与Lys57的氢键稳定活性位点区域，与Asp26合作调节Cys32的质子化状态。比较序列分析显示，在亲酸Trxs的43号位置经常发现小的非极性残基，如Gly和ala，这表明该位点的结构灵活性可能有助于酸性适应。为了测试该位点的替换如何影响Trx的稳定性和功能，我们在43号位置生成了EcTrx突变体（D43G, D43A, D43S, D43N， D43L和D43E）。热移和氯化胍（GdmCl）诱导的展开实验显示，D43A和D43S的稳定性显著降低，而D43G保持中等稳定性，这可能是由于在嗜酸乙酰杆菌Trx中观察到的更紧密的n端包装。这三个突变体也表现出增加的构象灵活性，而D43N和D43L通过极性或疏水侧链赋予部分稳定性。相比之下，d43e -模拟祖先的glue恢复了氢键，增强了热稳定性，形成了最刚性的结构。除D43S外，大多数突变体在DTNB试验中保持催化活性，其活性为野生型的~ 50% %。总之，我们的研究结果表明，Asp43对于维持EcTrx的结构稳定性至关重要，并表明该位点增强但不是过度的灵活性有助于Trx适应酸性环境。

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引用次数: 0

Molecular insights into diosgenin's role in preventing protein aggregation in neurodegenerative diseases 薯蓣皂苷元在神经退行性疾病中预防蛋白质聚集作用的分子研究。

IF 2.3 4区生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et biophysica acta. Proteins and proteomics

Pub Date : 2025-11-19 DOI: 10.1016/j.bbapap.2025.141116

Nagma Parveen , Faizan Abul Qais , Mohammad Faheem

Neurodegenerative disorders (ND) such as Parkinson's and Alzheimer's progressively impair the nervous system, leading to cognitive deterioration and motor dysfunction. A primary factor in these diseases is the accumulation of misfolded protein aggregates, which interfere with cellular processes and ultimately result in neuronal death. Preventing the formation of these toxic aggregates has the potential to protect neurons and slow the advancement of disease. This study examined the impact of diosgenin on protein aggregation, utilizing human serum albumin (HSA) as model protein. Diosgenin reduced ThT fluorescence by 64.35 % and decreased turbidity by 62.61 %, indicating a notable suppression of protein aggregation. The % α-helix in HSA experienced a decline from 57.68 % to 8.82 %, but diosgenin treatment restored it to 43.89 %. Binding studies demonstrated that diosgenin interacts with HSA with −11.0 kcal/mol binding energy, facilitated by van der Waals, hydrophobic and hydrogen bonding interactions, and stability of HSA-diosgenin complex was also validated using molecular simulations. To further elucidate the aggregation inhibition mechanism by diosgenin, advanced molecular dynamics simulations were employed. Diosgenin increased the solvent accessibility of the HSA₂₈₂_–₂₉₂ oligomers, reduced β-sheet formation, and prevented H-bond interactions, key factors in aggregate formation. Molecular simulation of Aβ oligomers (Aβ₁₆_–₂₂) also showed the diosgenin prevents oligomerization and β-sheet formation. We show that diosgenin presents a promising alternative due to its ability to stabilize protein structures and inhibit protein aggregation, making it a potential therapeutic candidate for NDs. However, further experimental validation in animal models is necessary to confirm diosgenin's anti-aggregation effects, particularly of amyloid-forming proteins.

神经退行性疾病（ND）如帕金森氏症和阿尔茨海默氏症逐渐损害神经系统，导致认知退化和运动功能障碍。这些疾病的一个主要因素是错误折叠的蛋白质聚集体的积累，它干扰细胞过程并最终导致神经元死亡。阻止这些有毒聚集体的形成有可能保护神经元并减缓疾病的进展。本研究以人血清白蛋白（HSA）为模型蛋白，研究了薯蓣皂苷元对蛋白质聚集的影响。薯蓣皂苷元使ThT荧光降低64.35 %，浊度降低62.61 %，表明对蛋白质聚集有明显的抑制作用。α-螺旋从57.68 %下降到8.82 %，薯蓣皂苷元处理后恢复到43.89 %。结合研究表明，薯蓣皂苷元与HSA相互作用的结合能为-11.0 kcal/mol，通过范德华、疏水和氢键相互作用，并通过分子模拟验证了HSA-薯蓣皂苷元配合物的稳定性。为了进一步阐明薯蓣皂苷元的聚集抑制机制，采用先进的分子动力学模拟方法。diogenin提高了HSA282-292低聚物的溶剂亲和性，减少了β-sheet的形成，并阻止了h -键相互作用，这是聚集体形成的关键因素。对Aβ低聚物（Aβ16-22）的分子模拟也表明薯蓣皂苷元可以抑制α β低聚和β-薄片的形成。我们发现薯蓣皂苷元由于其稳定蛋白质结构和抑制蛋白质聚集的能力而提供了一个很有希望的替代方案，使其成为nd的潜在治疗候选者。然而，进一步的动物模型实验验证是必要的，以确认薯蓣皂苷元的抗聚集作用，特别是淀粉样蛋白形成。

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引用次数: 0

Enhancement of the nucleotide incorporation activity of the terminal deoxynucleotidyl transferase by N-terminal truncation and DNA-binding protein modulation 通过n端截断和dna结合蛋白调节增强末端脱氧核苷酸转移酶的核苷酸结合活性。

IF 2.3 4区生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et biophysica acta. Proteins and proteomics

Pub Date : 2025-11-12 DOI: 10.1016/j.bbapap.2025.141114

Antos B. Sachanka, Stafaniya S. Douhaya, Veronika V. Shchur, Aliaksei V. Yantsevich

Terminal deoxynucleotidyl transferase (TdT) is a unique DNA polymerase that catalyzes the template-independent addition of nucleotides to the 3′ terminus of single-stranded DNA. Its distinctive catalytic properties have been exploited in aptasensor design, nanomaterial synthesis, DNA mutagenesis, innovative data storage based on single-nucleotide DNA sequences, and enzymatic de novo DNA synthesis. However, the application of the enzyme is limited by its low expression yield, poor thermostability, and reduced nucleotide incorporation efficiency with substrates prone to forming secondary structures. To overcome these limitations, we engineered a bovine TdT variant truncated at the N-terminus by 148 residues (148_TrTdT), which demonstrates a twofold increase in expression yield, a 5 °C improvement in thermostability, and over 30 % enhancement in nucleotide incorporation efficiency and processivity compared to the wild-type enzyme. Moreover, we demonstrated that DNA-binding proteins enhance the nucleotide incorporation activity of native TdT (10-fold) and 148_TrTdT (2-fold) when substrates are prone to forming secondary structures. Collectively, the methodologies implemented in this study exhibit significant potential for enhancing the efficiency of enzymatic de novo DNA synthesis, thereby enabling the development of new bioengineering and biomedical applications.

末端脱氧核苷酸转移酶（TdT）是一种独特的DNA聚合酶，它催化核苷酸在单链DNA 3'端独立于模板的添加。其独特的催化特性已被用于配体传感器设计、纳米材料合成、DNA诱变、基于单核苷酸DNA序列的创新数据存储和酶促从头合成DNA。然而，该酶的应用受到其表达量低、热稳定性差以及与底物形成二级结构的核苷酸结合效率降低的限制。为了克服这些限制，我们设计了一个在n端被148个残基截断的牛TdT变体（148_TrTdT），与野生型酶相比，其表达量增加了两倍，热稳定性提高了5 °C，核苷酸结合效率和加工效率提高了30 %以上。此外，我们还证明，当底物易于形成二级结构时，dna结合蛋白可增强天然TdT（10倍）和148_TrTdT（2倍）的核苷酸结合活性。总的来说，本研究中实施的方法在提高酶促从头DNA合成的效率方面表现出巨大的潜力，从而使新的生物工程和生物医学应用得以发展。

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引用次数: 0

Integrated computational and biological assays to identify and validate L. donovani homoserine dehydrogenase inhibitors 综合计算和生物测定鉴定和验证L. donovani高丝氨酸脱氢酶抑制剂。

IF 2.3 4区生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et biophysica acta. Proteins and proteomics

Pub Date : 2025-11-11 DOI: 10.1016/j.bbapap.2025.141112

Shalu Verma , Shaswat Sengar , Abdul Basit Khan , Anil Kumar Shakya , J. Venkatesh Pratap

The Neglected Tropical Disease (NTD) Leishmaniasis continues to pose a serious global health burden affecting millions globally. Visceral leishmaniasis, one of three clinical forms, can be fatal, if left untreated. Current treatment regimen relies mostly on re-purposed drugs and is limited by toxicity, high cost and the emergence of drug resistance, highlighting the need for safer, targeted therapies. In this study, we characterized Leishmania donovani homoserine dehydrogenase (LdHSD), a crucial enzyme in the aspartate pathway. As no human homolog exists, it represents a promising target for selective therapeutic intervention. LdHSD was purified to homogeneity, characterized biophysically and its activity biochemically assayed. Docking of the substrate (L-homoserine) and the cofactor (NADP⁺) to the LdHSD homology model identified adjacent but discrete binding pockets. Structure-based virtual screening of the Maybridge Compound Library was subsequently performed and the top 10 substrate-site bound compounds were taken for experimental validation. Two of the three piperidine/piperazine scaffold compounds inhibited LdHSD activity by ∼90 %. These findings highlight the therapeutic potential of targeting LdHSD for the development of novel, selective anti-leishmanial agents and establish a strong foundation for future optimization of these identified inhibitors.

被忽视的热带病利什曼病继续构成严重的全球健康负担，影响全球数百万人。内脏利什曼病是三种临床形式之一，如果不及时治疗，可能会致命。目前的治疗方案主要依赖于重新利用的药物，受到毒性、高成本和耐药性出现的限制，突出表明需要更安全的靶向治疗。在这项研究中，我们鉴定了利什曼原虫同型丝氨酸脱氢酶（LdHSD），这是天冬氨酸途径中的一个关键酶。由于没有人类同源物存在，它代表了选择性治疗干预的一个有希望的目标。对LdHSD进行纯化至均匀性，进行生物物理表征，并进行生物化学活性测定。底物（l -高丝氨酸）和辅因子（NADP）对接到LdHSD同源模型，确定了相邻但离散的结合口袋。随后对Maybridge化合物库进行基于结构的虚拟筛选，并选取前10个底物-位点结合化合物进行实验验证。三种哌啶/哌嗪支架化合物中的两种抑制LdHSD活性~90 %。这些发现突出了以LdHSD为靶点开发新型、选择性抗利什曼药物的治疗潜力，并为这些已确定的抑制剂的未来优化奠定了坚实的基础。

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引用次数: 0

Integrative multi-omics machine learning reveals novel driver genes associations in lung adenocarcinoma 综合多组学机器学习揭示了肺腺癌中新的驱动基因关联。

IF 2.3 4区生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et biophysica acta. Proteins and proteomics

Pub Date : 2025-11-08 DOI: 10.1016/j.bbapap.2025.141113

Fei Yuan , FeiMing Huang , Xiaoyu Cao , Yu-Hang Zhang , KaiYan Feng , YuSheng Bao , Tao Huang , Yu-Dong Cai

Lung adenocarcinoma remains a major challenge in cancer research due to its complex molecular underpinnings. In this study, we developed an integrated machine learning framework to identify novel driver genes associated with lung adenocarcinoma by leveraging multi-omics data. We curated gene candidates from methylation, RNAseq, mutation, and miRNA levels, and mapped them onto a protein–protein interaction network from STRING to generate informative feature vectors using a node2vec method. Furthermore, they were also represented by GO and KEGG enrichment features. All features were then refined through a multi-step process, beginning with the Boruta algorithm for filtering and followed by the minimum redundancy maximum relevance method for ranking. An incremental feature selection strategy was employed to determine the optimal feature subsets, which were used to build predictive models with random forest and support vector machine classifiers. To address class imbalance, synthetic sampling was applied, and ten-fold cross-validation ensured model robustness. Consequently, we predicted 428, 105, 1039, and 1748 potential lung adenocarcinoma driver genes for RNAseq, methylation, mutation, and miRNA levels, respectively. Integrated analysis of overlapping gene sets further highlighted key candidates, including PQLC3, FAM192A, FAM83D, SPRED1, SFTPB, and TM4SF5, with high composite probability scores. Some identified genes may be the driver genes of lung adenocarcinoma and have some druggable potential. These findings provide new insights into the molecular mechanisms of lung adenocarcinoma and suggest promising targets for future diagnostic and therapeutic strategies.

由于其复杂的分子基础，肺腺癌仍然是癌症研究的主要挑战。在这项研究中，我们开发了一个集成的机器学习框架，通过利用多组学数据来识别与肺腺癌相关的新驱动基因。我们筛选了来自甲基化、RNAseq、突变和miRNA水平的候选基因，并将它们映射到来自STRING的蛋白质-蛋白质相互作用网络上，使用node2vec方法生成信息丰富的特征向量。此外，它们还具有GO和KEGG富集特征。然后通过多步骤过程对所有特征进行细化，首先使用Boruta算法进行过滤，然后使用最小冗余最大相关性方法进行排序。采用增量特征选择策略确定最优特征子集，并结合随机森林和支持向量机分类器构建预测模型。为了解决类别不平衡问题，采用了合成抽样，并进行了十倍交叉验证，确保了模型的稳健性。因此，我们分别预测了428、105、1039和1748个潜在的肺腺癌驱动基因的RNAseq、甲基化、突变和miRNA水平。对重叠基因集的综合分析进一步突出了关键候选基因，包括PQLC3、FAM192A、FAM83D、SPRED1、SFTPB和TM4SF5，它们的复合概率得分很高。一些已鉴定的基因可能是肺腺癌的驱动基因，具有一定的药物潜力。这些发现为肺腺癌的分子机制提供了新的见解，并为未来的诊断和治疗策略提供了有希望的靶点。

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引用次数: 0

Novel interactions between FOXP2 and PAX6: Implications for neural development and autism spectrum disorders FOXP2和PAX6之间新的相互作用：对神经发育和自闭症谱系障碍的影响。

IF 2.3 4区生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et biophysica acta. Proteins and proteomics

Pub Date : 2025-11-08 DOI: 10.1016/j.bbapap.2025.141110

Jessica Brothwell , Olamide Jeje , Dineo Mashamaite , Sylvia Fanucchi

FOXP2 and PAX6 are transcription factors essential for neural development, with mutations in both linked to autism spectrum disorders (ASDs). Their DNA-binding domains include a forkhead domain (FHD) for FOXP2 and a paired domain (PD) plus homeodomain (HD) for PAX6. We investigated whether the FOXP2 FHD interacts directly with PAX6 PD or HD, and how such interactions influence DNA binding. Fluorescence anisotropy showed that all three domains bind specifically to their respective DNA targets with similar affinities. The FOXP2 FHD also interacts directly with both PAX6 PD and HD, with low micromolar binding affinities. Despite its stronger intrinsic DNA affinity, the FHD was displaced from its target DNA by both PAX6 domains, suggesting that protein–protein interactions can override DNA affinity under competitive conditions. In contrast, FOXP2 could not displace PD or HD from their DNA targets. Molecular docking supported these findings: DNA–protein interfaces were largely unchanged by the second protein, but protein–protein interfaces were strongly influenced by DNA occupancy. The H3 helix of FHD was identified as a central point for assembly, contributing to both DNA and protein interfaces. When FHD was bound to DNA, H3 was occupied, forcing PD or HD to dock at alternative, less optimal sites. HD maintained stronger contacts in these rearranged states, consistent with its greater competitive strength. This asymmetric interplay indicates competitive dominance by PAX6 and suggests mechanisms that could underlie transcriptional regulation in neurodevelopment.

FOXP2和PAX6是神经发育所必需的转录因子，两者的突变都与自闭症谱系障碍（asd）有关。它们的dna结合域包括FOXP2的叉头结构域（FHD）和PAX6的成对结构域（PD）加同源结构域（HD）。我们研究了FOXP2 FHD是否直接与PAX6 PD或HD相互作用，以及这种相互作用如何影响DNA结合。荧光各向异性表明，这三个结构域特异性结合各自的DNA目标具有相似的亲和力。FOXP2 FHD也直接与PAX6 PD和HD相互作用，具有低微摩尔结合亲和力。尽管FHD具有更强的内在DNA亲和力，但它被两个PAX6结构域从目标DNA中取代，这表明在竞争条件下，蛋白质-蛋白质相互作用可以覆盖DNA亲和力。相比之下，FOXP2不能将PD或HD从它们的DNA靶标上取代。分子对接支持这些发现：DNA-蛋白质界面在很大程度上不受第二个蛋白质的影响，但蛋白质-蛋白质界面受到DNA占用的强烈影响。FHD的H3螺旋被确定为组装的中心点，有助于DNA和蛋白质的界面。当FHD与DNA结合时，H3被占用，迫使PD或HD停靠在替代的、不太理想的位置。HD在这些重组后的州保持了更强的联系，这与它更强的竞争实力相一致。这种不对称的相互作用表明PAX6具有竞争性优势，并提示了神经发育中转录调节的机制。

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引用次数: 0

Biophysical characterization of Hpa1 protein from Xanthomonas orzyae 黄单胞菌Hpa1蛋白的生物物理特性研究。

IF 2.3 4区生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et biophysica acta. Proteins and proteomics

Pub Date : 2025-11-07 DOI: 10.1016/j.bbapap.2025.141111

Jaimini Patoliya , Khushali Thaker , Jahanvi Bajpai , Dhaval Patel , Nayan Jain , Prasant Kumar , Rushikesh Joshi

Xanthomonas oryzae pv. oryzae (Xoo), the causative agent of bacterial blight in rice, secretes a suite of effectors via the Type III Secretion System (T3SS), among which harpin proteins like Hpa1 are key modulators of plant immunity. Unlike other T3SS effectors, Hpa1 lacks enzymatic activity and instead acts as a biostimulant, promoting growth and defense responses. In this study, the hpa1 gene was cloned from Xoo BXO1 strain and successfully expressed in E. coli Rosetta cells. The recombinant protein was purified using Ni-NTA chromatography, and SDS-PAGE, Western blotting, and MALDI-TOF analysis confirmed its molecular weight (∼16.5 kDa). Biophysical characterization revealed a predominance of α-helical content, with CD spectroscopy. Thermal and chemical unfolding of Hpa1 was evaluated using CD spectroscopy (with GuHCl) and ANS fluorescence (with GuHCl and urea), revealing moderate stability. pH-dependent stability assessed by fluorescence showed optimal structural retention at pH 5. TFE-induced structural modulation, studied by CD, demonstrated concentration-dependent α-helix enhancement. Thermal stability was assessed through four approaches: SDS-PAGE, CD at 222 nm, SYPRO Orange-based thermal shift assay, and HR-inducing activity in Nicotiana benthamiana. These results collectively confirmed that native Hpa1 conformation is not necessary for its function. These findings highlight its potential as a protein-based biostimulant for agricultural applications and its utility as a model for studying intrinsically disordered yet functionally stable proteins.

米黄单胞菌。水稻白叶枯病病原菌oryzae （Xoo）通过III型分泌系统（Type III分泌系统，T3SS）分泌一系列效应物，其中Hpa1等harpin蛋白是植物免疫的关键调节剂。与其他T3SS效应物不同，Hpa1缺乏酶活性，而是作为生物刺激物，促进生长和防御反应。本研究从Xoo BXO1菌株中克隆了hpa1基因，并在大肠杆菌Rosetta细胞中成功表达。重组蛋白经Ni-NTA层析纯化，SDS-PAGE、Western blotting和MALDI-TOF分析证实其分子量为~16.5 kDa。生物物理表征表明α-螺旋含量占主导地位。利用CD光谱（含GuHCl）和ANS荧光（含GuHCl和尿素）对Hpa1的热展开和化学展开进行了评估，结果显示Hpa1的稳定性中等。荧光测定的pH依赖性稳定性表明，pH为 5时结构保持最佳。CD研究了tfe诱导的结构调制，显示出浓度依赖性α-螺旋增强。通过四种方法评估热稳定性：SDS-PAGE、222 nm CD、SYPRO橙基热移法和benthamiana诱导hr活性。这些结果共同证实了天然Hpa1构象对其功能不是必需的。这些发现突出了它作为一种基于蛋白质的生物刺激素在农业应用中的潜力，以及它作为研究内在无序但功能稳定的蛋白质的模型的效用。

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引用次数: 0

Self-association of Apolipoprotein A-I studied with multiple-association models: A pyrene multiparametric analysis. 多关联模型研究载脂蛋白A- i的自关联：芘多参数分析。

IF 2.3 4区生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et biophysica acta. Proteins and proteomics

Pub Date : 2025-11-01 Epub Date: 2025-08-06 DOI: 10.1016/j.bbapap.2025.141093

Wilson A Tárraga, Lisandro J Falomir-Lockhart, Horacio A Garda, Marina C Gonzalez

Apolipoprotein A-I (apoA-I) is the main protein of high-density lipoprotein particles, conferring anti-atherogenic properties through reverse cholesterol transport. In its lipid-free state, apoA-I self-associates, a process implicated in normal physiology as well as in pathological conditions, such as amyloidosis. Although various mechanisms have been proposed to explain its self-association, there is still no consensus, and its functional implications remain unclear. Here, we employed a multi-parametric fluorescent probe to investigate apoA-I self-association. We used three single cysteine mutants located in different helixes: K107C (H4), K133C (H5), and F225C (H10); and labelled them with pyrene as detailed previously (Tárraga et al. Arch Biochem Biophys 699 (2021) 108748). Our original experiments revealed excimer emission between helixes H5 and H10 and polarity changes also in H4. By exploiting specific pyrene band emissions to monitor dimer association (via excimer formation) and microenvironment polarity (P-value), we tracked apoA-I oligomerization as a function of protein concentration. Several mathematical models of self-association were developed and compared using selection criteria to identify the simplest model reproducing apoA-I's complex behaviour in aqueous media: a Sequential Association submodel limited to a tetramer as the highest order oligomeric species and considering both dimers and tetramers as responsible for the excimer's emission. Multi-equilibria models enhanced the titration analysis, allowing estimation of association constants (Ka) and oligomeric species distribution. Our results support previous evidence that contacts among helixes, which stabilize discoidal HDL particles, are already present in lipid-free apoA-I, with dimeric species predominating, and possible tetrameric too. Further investigation of these species is essential to elucidate their physiological and pathological roles, such as in atherosclerosis.

载脂蛋白A-I （apoA-I）是高密度脂蛋白颗粒的主要蛋白，通过逆向胆固醇转运具有抗动脉粥样硬化特性。在无脂状态下，apoa - 1自我结合，这一过程涉及正常生理和病理条件，如淀粉样变性。尽管人们提出了各种机制来解释其自我关联，但仍未达成共识，其功能含义仍不清楚。在这里，我们采用多参数荧光探针来研究apoA-I的自关联。我们使用了三个位于不同螺旋上的单半胱氨酸突变体：K107C （H4）、K133C （H5）和F225C (H10)；如前所述，用芘标记它们（Tárraga等）。生物化学学报，699(2021)108748。我们最初的实验发现H5和H10螺旋之间的准分子发射和H4的极性也发生了变化。通过利用特定的芘带发射来监测二聚体结合（通过准分子形成）和微环境极性（p值），我们跟踪了apoA-I寡聚化作为蛋白质浓度的函数。研究人员开发了几种自结合的数学模型，并使用选择标准进行了比较，以确定再现apoA-I在水介质中复杂行为的最简单模型：一个顺序结合子模型，该模型将四聚体作为最高阶寡聚物，并考虑到二聚体和四聚体都负责准分子的发射。多平衡模型增强了滴定分析，允许估计缔合常数（Ka）和低聚物种分布。我们的研究结果支持了先前的证据，即在无脂apoa - 1中已经存在稳定盘状HDL颗粒的螺旋之间的接触，二聚体占主导地位，也可能是四聚体。进一步研究这些物种对于阐明其生理和病理作用至关重要，例如动脉粥样硬化。

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引用次数: 0

Drug–biomolecular interaction: Spectroscopic and computational insights into esomeprazole binding with human serum albumin 药物-生物分子相互作用：埃索美拉唑与人血清白蛋白结合的光谱和计算见解。

IF 2.3 4区生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et biophysica acta. Proteins and proteomics

Pub Date : 2025-10-31 DOI: 10.1016/j.bbapap.2025.141109

Anna Tanuja Safala Bodapati , Ragaiahgari Srinivas Reddy , Kandikonda Lavanya , Shravya Rao Madku , Rajdeep Chowdhury , Bijaya Ketan Sahoo

Cellular process relies on specific binding of drug molecules with desired biological receptors. Biomolecular receptors and drugs exhibit their biological functions upon their interactions. Understanding the binding parameters and energetics of the binding provides a plethora of information that may be helpful in drug design and discovery. Further, drug interaction with carrier proteins affects the pharmacokinetics and dynamics of other exogenous and endogenous drugs. In this work, we report the binding behavior of esomeprazole with human serum albumin using several biophysical techniques. Qualitative and quantitative aspects of the binding along with the binding energetics has been focused in extracting the thermodynamics parameters and key interaction force. The binding constants (K_b)obtained from Scatchard analysis are-1.17 × 10⁵, 7.49 × 10⁴, and 3.23 × 10⁴ M⁻¹ at 298, 303, 308 K respectively. Esomeprazole binds preferentially at site 1 in subdomain IIA of human serum albumin and was confirmed, supported by site-specific marker displacement studies and docking simulations. Negative value of change in free energy (∆G⁰) -32 kJ/mol at 298 K showed thermodynamically favourable interaction and outweigh of entropic factor (T∆S⁰ = 230.76 ± 3 kJ for T = 298 K) over the enthalpic contribution (∆H⁰ = −261.99 kJ/mol) revealed an entropy-driven process. Binding of esomeprazole affected the helical structure of human serum albumin. Molecular docking studies (theoretical) shows the binding pocket of ESM at site1(IIA), which is in accordance with the experimental result. Further, the interface residues involved in the binding were analysed from the 2D diagram and ligplot of the docked complex.

细胞过程依赖于药物分子与所需生物受体的特异性结合。生物分子受体与药物通过相互作用发挥其生物学功能。了解结合参数和结合的能量学提供了大量的信息，可能有助于药物设计和发现。此外，药物与载体蛋白的相互作用会影响其他外源性和内源性药物的药代动力学和动力学。在这项工作中，我们使用几种生物物理技术报道了埃索美拉唑与人血清白蛋白的结合行为。结合热力学参数和关键作用力的提取，从定性和定量两个方面进行了研究。Scatchard分析得到的结合常数（Kb）在298、303、308 K处分别为1.17 × 105、7.49 × 104和3.23 × 104 M-1。埃索美拉唑优先结合人血清白蛋白IIA亚结构域的1号位点，这一发现得到了位点特异性标记位移研究和对接模拟的证实。负值自由能的变化(∆G0) -32 焦每摩尔298 K显示热力学有利的互动和大于熵的因子(T∆S0 = 230.76 ± 3 kJ 298 T =  K)在向左反应贡献(∆H0 = -261.99 焦每摩尔)显示一个熵驱动过程。埃索美拉唑的结合影响了人血清白蛋白的螺旋结构。分子对接研究（理论）显示ESM的结合袋位于site1(IIA)，与实验结果一致。此外，通过对接配合物的二维图和光图分析了参与结合的界面残基。

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引用次数: 0

Multifunctionality analysis of serine hydroxymethyltransferases from human and Escherichia coli 人和大肠杆菌丝氨酸羟甲基转移酶的多功能分析

IF 2.3 4区生物学 Q3 BIOCHEMISTRY & MOLECULAR BIOLOGY

Biochimica et biophysica acta. Proteins and proteomics

Pub Date : 2025-10-25 DOI: 10.1016/j.bbapap.2025.141107

Mahiro Hayashi , Kumiko Sakai-Kato , Tetsuya Miyamoto

Serine hydroxymethyltransferase (SHMT) catalyzes the conversion of l-serine and tetrahydrofolate (THF) to glycine and 5,10-methylenetetrahydrofolate, respectively. In a previous study, we found that human and Escherichia coli SHMTs possess THF-dependent d-serine dehydratase activity, which degrades d-serine to pyruvate and ammonia. Some activities including aldolase and racemase activities have also been reported for SHMTs. In the present study, we investigated the multifunctionality of E. coli SHMT and two human SHMTs. All three SHMTs displayed high aldolase activity toward l-allo-threonine and l-threo-phenylserine, and measurable activity toward l-threonine, but they did not act on d-allo-threonine and d-threonine. The catalytic efficiency (k_cat/K_m) for l-allo-threonine was higher than for l-threo-phenylserine. None of the SHMTs displayed racemase activity toward various amino acids, although slight alanine racemase activity was detected for E. coli SHMT. Likewise, none of the SHMTs showed lyase, aminotransferase, or aspartate decarboxylase activities, and none exhibited dehydratase activity toward other hydroxy amino acids except d-serine. SHMTs are multifunctional enzymes possessing canonical hydroxymethyltransferase, d-serine dehydratase, and low-specificity l-threonine aldolase activities.

丝氨酸羟甲基转移酶（SHMT）催化l-丝氨酸和四氢叶酸（THF）分别转化为甘氨酸和5,10-亚甲基四氢叶酸。在之前的研究中，我们发现人类和大肠杆菌的SHMTs具有thf依赖性d-丝氨酸脱水酶活性，可将d-丝氨酸降解为丙酮酸和氨。一些活性，包括醛缩酶和消旋酶活性也被报道为SHMTs。在本研究中，我们研究了大肠杆菌SHMT和两种人类SHMT的多功能性。所有三种shmt对l-异体苏氨酸和l-三苯基丝氨酸均表现出较高的醛缩酶活性，对l-苏氨酸具有可测量的活性，但对d-异体苏氨酸和d-苏氨酸没有作用。l-异丙苏氨酸的催化效率（kcat/Km）高于l-三苯基丝氨酸。虽然在大肠杆菌SHMT中检测到轻微的丙氨酸外消旋酶活性，但没有一种SHMT显示出对各种氨基酸的外消旋酶活性。同样，没有一个SHMTs显示裂解酶、转氨酶或天冬氨酸脱羧酶活性，也没有显示除d-丝氨酸外的其他羟基氨基酸的脱水酶活性。SHMTs是多功能酶，具有典型的羟甲基转移酶、d-丝氨酸脱水酶和低特异性的l-苏氨酸醛缩酶活性。

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引用次数: 0