Animals
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2025
·Vol. 15
·期 6
·865
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IF 2.7
Chengcheng Wu,Lihan Tao,Quanyong Zhou,Fanfan Zhang,Yanbing Zeng
Simple SummaryThe helicase NSP13, encoded by porcine deltacoronavirus (PDCoV), regulates viral replication and is considered to bean ideal target for antiviral drugs. In our work, the wild-type NSP13WTand various deletion mutants were generated through expression and purification, and we tested the activities of these proteins using multiple methods. The results revealed that both the Stalk and 1B domains were necessary for NSP13WTto separate the double-stranded nucleic acids, and the deletion of the ZBD impaired this unwinding activity. Our study is the first to explore the role of each domain in the activities of PDCoV NSP13, providing mechanistic insights into helicase-regulated viral replication and laying the foundation for the development of antiviral drugs.The nonstructural protein 13 (NSP13) of PDCoV is a highly conservative helicase and plays key roles in viral replication. NSP13 contains a zinc-binding domain (ZBD), a helical Stalk domain, a beta-barrel 1B domain, and a core helicase domain. However, the specific functions of these domains of PDCoV NSP13 remain largely unknown. Here, we expressed and purified the wild-type NSP13WTand various mutants with domain deletions, and the activities of these proteins were analyzed using multiple methods. We found that NSP13ΔZBDpossessed the abilities to hydrolyze ATP and unwind double-stranded nucleic acids, but the unwinding efficiency was lower than that of NSP13WT. In contrast, NSP13ΔZBD-Stalk, NSP13Δ1B, and NSP13ΔZBD-Stalk-1Ball lost their unwinding activity, but not their ATPase activity. These results revealed that the deletion of the ZBD impaired the unwinding activity of PDCoV helicase NSP13, and the Stalk and 1B domains were critical for NSP13 to separate the duplexes. The identification of the roles of each domain in this study was helpful to gain an in-depth understanding of the overall functions of helicase NSP13, providing a theoretical basis for the development of antiviral drugs targeting helicase.
研究领域:
病毒学
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分子生物学
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抗病毒药物开发
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DOI
PubMed
高影响因子
EMBO JOURNAL
·
2020
·Vol. 39
·期 16
·e103631
·
IF 9.889
Xianping Wang,Jihong Gong,Le Zhu,Shen Wang,Xiaoyu Yang,Yuanyuan Xu,Xiaofei Yang,Cong Ma
Priming of synaptic vesicles involves Munc13-catalyzed transition of the Munc18-1/syntaxin-1 complex to the SNARE complex in the presence of SNAP-25 and synaptobrevin-2; Munc13 drives opening of syntaxin-1 via the MUN domain while Munc18-1 primes SNARE assembly via domain 3a. However, the underlying mechanism remains unclear. In this study, we have identified a number of residues in domain 3a of Munc18-1 that are crucial for Munc13 and Munc18-1 actions in SNARE complex assembly and synaptic vesicle priming. Our results showed that two residues (Q301/K308) at the side of domain 3a mediate the interaction between the Munc18-1/syntaxin-1 complex and the MUN domain. This interaction enables the MUN domain to drive the opening of syntaxin-1 linker region, thereby leading to the extension of domain 3a and promoting synaptobrevin-2 binding. In addition, we identified two residues (K332/K333) at the bottom of domain 3a that mediate the interaction between Munc18-1 and the SNARE motif of syntaxin-1. This interaction ensures Munc18-1 to persistently associate with syntaxin-1 during the conformational change of syntaxin-1 from closed to open, which reinforces the role of Munc18-1 in templating SNARE assembly. Taken together, our data suggest a mechanism by which Munc13 activates the Munc18-1/syntaxin-1 complex and enables Munc18-1 to prime SNARE assembly.
研究领域:
神经科学
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细胞生物学
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分子生物学
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DOI
PubMed
高影响因子
Journal of Translational Medicine
·
2025
·Vol. 23
·期 1
·1-18
·
IF 7.5
Tan Yanzhen,Li Hong,Cao Guojie,Xin Jialin,Yan Dongxu,Liu Yingying,Li Panpan,Zhang Yuxi,Shi Lei,Zhang Bing,Yi Wei,Sun Yang
Background In developed nations, myocardial infarction (MI) is one of the main causes of morbidity and mortality, resulting in a significant economic burden and becoming a global public health problem. C1q/tumor necrosis factor-related protein 9 (CTRP9) is a secreted protein comprising a variable domain, a collagenous region, and a C-terminal trimerizing globular C1q (gC1q) domain. In vivo, the full-length CTRP9 (fCTRP9) can be cleaved into the globular domain of CTRP9 (gCTRP9). Here, we tested the cardio-protective impacts of fCTRP9, gCTRP9, and N-terminal domain, including the variable and collagenous domain, of CTRP9 (nCTRP9) in the context of MI.Methods Studies comparing the protective properties of fCTRP9 and gCTRP9 against MI in mice hearts were performed both in vitro and in vivo. The role of matrix metalloproteinase-9 (MMP9) in CTRP9 cleavage was examined, and the effects of different CTRP9 domains on cardiac fibrosis and cardiac fibroblast (CF) activation were investigated.Results gCTRP9 exerted better protective effects than fCTRP9 against MI, demonstrating superior anti-apoptotic and anti-fibrotic properties. fCTRP9 was cleaved by MMP9, resulting in gCTRP9 and nCTRP9. MMP9 overexpression enhanced the cardioprotective effects of fCTRP9, while nCTRP9 supplementation aggravated cardiac fibrosis in MI mice. Mechanistically, nCTRP9 activated CFs via an increase in Rap1 expression and MEK 1/2 and ERK1/2 phosphorylation.Conclusion sDifferent domains of CTRP9 have distinct cardioprotective effects. gCTRP9 shows beneficial effects, while nCTRP9 promotes cardiac fibrosis. These findings highlight the importance of CTRP9 in cardiac function regulation and suggest prospective therapeutic options for MI treatment.
研究领域:
心脏病学
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分子生物学
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药理学
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DOI
PubMed
高影响因子
MOLECULAR PHARMACEUTICS
·
2024
·
IF 4.5
Wanjun Han,Zheyue Bo,Ting Liang,Han Liu,Lanjing Li,Zhening Guo,Ru Huan,Bruno Hagenbuch,Chunshan Gui
Organic anion-transporting polypeptides (OATPs) 1B1 and 1B3 are two highly homologous transport proteins. However, OATP1B1- and 1B3-mediated estradiol-17β-glucuronide (E17βG) uptake can be differentially affected by clotrimazole. In this study, by functional characterization on chimeric transporters and single mutants, we find that G45 in transmembrane domain 1 (TM1) and V386 in TM8 are critical for the activation of OATP1B3-mediated E17βG uptake by clotrimazole. However, the effect of clotrimazole on the function of OATP1B3 is substrate-dependent as clotrimazole does not stimulate OATP1B3-mediated uptake of 4′,5′-dibromofluorescein (DBF) and rosuvastatin. In addition, clotrimazole is not transported by OATP1B3, but it can efficiently permeate the plasma membrane due to its lipophilic properties. Homology modeling and molecular docking indicate that E17βG binds in a substrate binding pocket of OATP1B3 through hydrogen bonding and hydrophobic interactions, among which its sterol scaffold forms hydrophobic contacts with V386. In addition, a flexible glycine residue at position 45 is essential for the activation of OATP1B3. Finally, clotrimazole is predicted to bind at an allosteric site, which mainly consists of hydrophobic residues located at the cytoplasmic halves of TMs 4, 5, 10, and 11.
研究领域:
药理学
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分子生物学
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DOI
PubMed
高影响因子
DEVELOPMENTAL AND COMPARATIVE IMMUNOLOGY
·
2019
·
IF 3.119
Liting Wu,Enxu Zhou,Along Gao,Linghe Kong,Siwei Wu,Xia Bian,Yuan Li,Bingxi Li,Shengli Fu,Zheng Guo,Jianmin Ye
B lymphocyte-induced maturation protein 1 (Blimp-1), a transcription factor containing zinc finger , is required and sufficient to trigger terminal differentiation of B cells in mammals. The Blimp-1 ( OnBlimp-1 ) from Nile tilapia ( Oreochromis niloticus ) was identified and characterized its expression pattern during B cell activation and maturation. The cDNA of OnBlimp-1 open reading frame is 2547 bp encoding a protein of 848 amino acids and the predicted molecular weight is 93.36 kDa. OnBlimp-1 contains a SET domain and five ZnF_C2H2 domains, which shares high homology with that of other species. OnBlimp-1 transcription was detected in all examined tissues with high expression in the spleen (SPL). Analysis of sorted lymphocyte populations , including IgM + and IgM − cells from peripheral blood (PBL), SPL and anterior kidney (AK), indicated that the OnBlimp-1 transcription was highly expressed in the IgM + B cells. Upon LPS stimulation, OnBlimp-1 expression was up-regulated in tissues of PBL, SPL and AK significantly. The expression of OnBlimp-1 , as well as the secreted IgM , was significantly up-regulated in the SPL and AK leukocytes stimulated with anti-OnIgM monoclonal antibody and LPS in vitro , respectively. Above results suggest that OnBlimp-1, a cytokine regulating the terminal differentiation of activated B cells to antibody-secreting cells, is likely to play important roles in B cell activation and maturation in Nile tilapia.
研究领域:
免疫学
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分子生物学
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鱼类生理学
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DOI
PubMed
高影响因子
IN VITRO CELLULAR & DEVELOPMENTAL BIOLOGY-ANIMAL
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2025
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IF 1.7
Qiu Yu,Ruan Huijie,Ji Decai,Cao Na
Bladder cancer is a common malignancy in the genitourinary system with its incidence rate among the world's highest. Neuralized E3 ubiquitin protein ligase 1 (NEURL1) belongs to the RING E3 ubiquitin ligase family, and its role in bladder cancer has not been reported yet. We aimed to explore the expression and roles of NEURL1 in bladder cancer. The NEURL1 expression was determined in clinical samples of bladder cancer. We stably overexpressed NEURL1 and NEURL1 with the RING domain deletion in human bladder cancer cell lines 5637 and RT-112 to investigate its functions. NEURL1 was confirmed to be significantly down-regulated in clinical bladder tumor specimens. NEURL1 overexpression significantly inhibited the growth, colony formation, and Ki-67 protein expression in both bladder cancer cells. The overexpression of NEURL1 also increased the apoptosis rate and cleaved caspase-3 protein expression in 5637 and RT-112 cells. As expected, the RING-deleted NEURL1 had no such effect. Moreover, NEURL1 overexpression facilitated the apoptosis of 5637/RT-112 cells under cisplatin conditions. NEURL1 promotes ubiquitination and proteasomal degradation of PDE9A. PDE9A protein expression was notably increased in bladder tumors from clinical specimens. Overexpressed NEURL1 inhibited PDE9A protein expression in both cell lines, whereas NEURL1 with the RING domain deletion did not. The addition of proteasome inhibitors MG-132 reversed the decrease in PDE9A expression by NEURL1 overexpression. Cell viability was inhibited and apoptosis was increased in the 5637 and RT-112 cells with stable knockdown of PDE9A. NEURL1 may be involved in the bladder cancer progression by increasing apoptosis, thereby leading to tumor cell growth inhibition.
研究领域:
肿瘤学
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分子生物学
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泌尿外科
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DOI
PubMed
高影响因子
FEBS LETTERS
·
2018
·Vol. 593
·期 2
·175-186
·
IF 2.999
Shengnan Yang,Yuan Zhang,Fukui Shen,Xiaoyao Ma,Man Zhang,Yuanyuan Hou,Gang Bai
Baicalin is one of the main flavonoids of the dried root of Scutellaria baicalensis Georgi and is reported to exert beneficial effects on the regulation of glucose/lipid metabolism. However, understanding its specific target and unique mechanism for improving glucose utilization is a challenge. In this paper, target fishing with a baicalin probe reveals that baicalin interacts with AKT. An immunofluorescence assay further demonstrates the colocalization of baicalin with AKT in the cytoplasm. A competitive test and virtual docking show that baicalin might bind to the pleckstrin homology domain of AKT. This specific binding hampers AKT membrane translocation, activates the phosphorylation of AKT on Ser473, induces the downstream glycogen synthase kinase 3β activation, and affects glycogen synthesis.
研究领域:
药理学
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分子生物学
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代谢
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DOI
PubMed
高影响因子
INTERNATIONAL JOURNAL OF BIOLOGICAL MACROMOLECULES
·
2024
·
IF 7.7
Hao Liu,Min Huang,Shuxin Wei,Xiaowen Wang,Yaqin Zhao,Zhengyang Han,Xianfeng Ye,Zhoukun Li,Yanling Ji,Zhongli Cui,Yan Huang
β-1,3-Galactanases selectively degrade β-1,3-galactan, thus it is an attractive enzyme technique to map high-galactan structure and prepare galactooligosaccharides. In this work, a gene encoding exo -β-1,3-galactanase (PxGal43) was screened form Paenibacillus xylanexedens , consisting of a GH43 domain, a CBM32 domain and α-L-arabinofuranosidase B (AbfB) domain. Using β-1,3-galactan (AG-II-P) as substrate, the recombined enzyme expressed in Escherichia coli BL21 (DE3) exhibited an optimal activity at pH 7.0 and 30 °C. The enzyme was thermostable, retaining >70 % activity after incubating at 50 °C for 2 h. In addition, it showed high tolerance to various metal ions, denaturants and detergents. Substrate specificity indicated that PxGal43 hydrolysis only β-1,3-linked galactosyl oligosaccharides and polysaccharides, releasing galactose as an exo -acting manner. The function of the CBM32 and AbfB domain was revealed by their sequential deletion and suggested that their connection to the catalytic domain was crucial for the oligomerization, catalytic activity, substrate binding and thermal stability of PxGal43. The substrate docking and site-directed mutagenesis proposed that Glu191, Gln244, Asp138 and Glu81 served as the catalytic acid, catalytic base, pKa modulator, and substrate identifier in PxGal43, respectively. These results provide a better understanding and optimization of multi-domain bacterial GH43 β-1,3-galactanase for the degradation of arabinogalactan.
研究领域:
生物化学
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酶学
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微生物学
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DOI
PubMed
高影响因子
BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS
·
2018
·
IF 2.559
Peihai Li,Kun Zang,Yingjie Li,Changshui Liu,Qingjun Ma
Extracellular proteases are often produced as pre-pro-enzyme and then undergo multiple processing steps to mature into the active form. The protease Epp, a virulent factor of Vibrio anguillarum , belongs to this family. Its maturation might be regulated by Ca 2+ via its polycystic kidney disease (PKD) domain, but the molecular mechanism is unknown. Herein, we report the crystal structure of the first PKD domain from V. anguillarum Epp (Epp-PKD1) and its specific Ca 2+ -binding capacity. Epp-PKD1 exists as a monomer , consisting of seven β-strands which form two β-sheets stacking with each other. One Ca 2+ is bound by the residues Asn3, Gln4, Asp27, Asp29, Asp68 and a water molecule with a pentagonal bipyramidal geometry. Incubating the apo Epp-PKD1 with Ca 2+ but not Mg 2+ , Mn 2+ , or Zn 2+ , enhances the thermal and chemical stability of Epp-PKD1, indicating its specific binding to Ca 2+ . Epp-PKD1 shares high similarity in both sequence and overall structure with that of Vibrio cholerae PrtV, a homologous protease of Epp, however, they differ in the oligomeric state and local structure at the Ca 2+ -binding site, suggesting maturation of PrtV and Epp might be differently regulated by Ca 2+ . Likely, proteases may take advantage of the structural diversity in PKD domains to tune their Ca 2+ -regulated maturation process.
研究领域:
生物化学
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结构生物学
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微生物学
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DOI
高影响因子
BRAIN RESEARCH
·
2025
·
IF 2.6
Xingli Tan,Xiaoli Su,Ying Wang,Weiwei Liang,Di Wang,Di Huo,Hongyong Wang,Yan Qi,Wenmo Zhang,Ling Han,Dongmei Zhang,Ming Wang,Jing Xu,Shuyu Wang,Jing Wang,Honglin Feng
Dysregulation of copper and iron homeostasis contributes to the progression of amyotrophic lateral sclerosis (ALS), but the role and mechanism of COMM domain containing 4 (COMMD4) in ALS remains unclear. In this research, we showed that the expression of COMMD4 was upregulated in ALS cells and animal models. The increased COMMD4 induced neuronal ferroptosis by disrupting the Cu-Fe balance. Mechanistic studies indicated that COMMD4 inhibited ferroportin (FPN)-mediated neuronal iron efflux by inhibiting intracellular copper and hephaestin (HEPH). Our findings demonstrated that COMMD4 depletion exerts neuroprotective effects on ALS by increasing intracellular copper and activating HEPH/FPN pathway, rather than affecting the interaction between HEPH and FPN. Targeting COMMD4 and its downstream signaling pathways may offer potential therapeutic avenues for ALS.
研究领域:
神经生物学
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分子生物学
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医学
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产品关联
DOI
PubMed
高影响因子
Insect Science
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2025
·
IF 3
Junna Liu,Zhaoyang Li,Zexin Zhong,Jianxing Li,Qi Chen,Ran Gao,Xuguo Zhou,Youjun Zhang,Chunxiao Yang,Huipeng Pan
LIM-HD (homeodomain) protein is essential for the specific differentiation of various cells and tissues due to its conserved HD. Within the LIM subclass, the LIM-HD transcription factor 1 alpha ( Lmx1a ) remains poorly characterized in insects. This study investigates the expression patterns of HvLmx1a in the 28-spotted lady beetle, Henosepilachna vigintioctopunctata , at both developmental stage and tissue levels. Furthermore, RNA interference (RNAi)-mediated knockdown of HvLmx1a resulted in increased mortality during the early larval stage. Injection of ds HvLmx1a at the beginning of the 4th instar caused a reduction in 20E titer, disrupted normal molting and pupation processes, and led to the emergence of larval-pupal abnormalities. Notably, HvLmx1a expression was significantly down-regulated on d 2 and 4 post-injection, which coincided with a marked inhibition of HvCHS1 and genes associated with 20E and bursicon signaling pathways. Additionally, silencing HvLmx1a induced qualitative changes in male testes and female ovaries, resulting in infertility and increased mortality. Ultimately, these findings suggest that HvLmx1a influences ovarian morphology and development through lipid metabolism. In conclusion, this study provides the first evidence of the diverse physiological roles of Lmx1a in H. vigintioctopunctata and highlight its potential as a target for RNAi-based biological control strategies.
研究领域:
昆虫学
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分子生物学
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发育生物学
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DOI
PubMed
高影响因子
Cell Reports
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2019
·
IF 7.815
Peilin Yu,Zhenming Liu,Xiafei Yu,Peiwu Ye,Huan Liu,Xiwen Xue,Lixin Yang,Zhongtang Li,Yang Wu,Cheng Fang,Yong Juan Zhao,Fan Yang,Jian Hong Luo,Lin-Hua Jiang,Liangren Zhang,Lihe Zhang,Wei Yang
Summary cADPR is a well-recognized signaling molecule by modulating the RyRs, but considerable debate exists regarding whether cADPR can bind to and gate the TRPM2 channel, which mediates oxidative stress signaling in diverse physiological and pathological processes. Here, we show that purified cADPR evoked TRPM2 channel currents in both whole-cell and cell-free single-channel recordings and specific binding of cADPR to the purified NUDT9-H domain of TRPM2 by surface plasmon resonance. Furthermore, by combining computational modeling with electrophysiological recordings, we show that the TRPM2 channels carrying point mutations at H1346, T1347, L1379, S1391, E1409, and L1484 possess distinct sensitivity profiles for ADPR and cADPR. These results clearly indicate cADPR is a bona fide activator at the TRPM2 channel and clearly delineate the structural basis for cADPR binding, which not only lead to a better understanding in the gating mechanism of TRPM2 channel but also shed light on a cADPR-induced RyRs-independent Ca 2+ signaling mechanism.
研究领域:
细胞生物学
·
分子生物学
·
生理学
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DOI
高影响因子
危险品化学品经营许可证（带存储）