搜索结果：Sheng wu gong cheng xue bao = Chinese journal of biotechnology

Global salinized land greatly affects the ecology and agriculture. Biological remediation is an important green approach for managing salinized soil. Enhancing the salt tolerance of plant growth-promoting rhizobacteria (PGPR) is key to improving the effectiveness of biological remediation. Glycine betaine (GB), an important osmotic regulator, can enhance the stress resistance of microorganisms and plants. In order to improve the survival rate of PGPR in salinized soil, GB was synthesized by biological method. In this study, we used a novel PGPR strain Bacillus paralicheniformis Bp1 as the host to reconstruct the GB synthesis pathway. First, the genes maeA, aceB, iclR, and gcvP in the glycine competition/consumption pathway were knocked out, and the glyoxylate cycle was strengthened by introduction of aceAK from Escherichia coli. Additionally, the exogenous high-efficiency transaminase gene agx1 was introduced. The highest yield of GB precursor, glycine, reached 34.27 mg/L within 30 h of fermentation, representing a 293.9% increase compared with the highest yield of the wild-type at the time point of 12 h. Meanwhile, to ensure the stability of the strain, aceAK was integrated into the genome to obtain strain Bp1Z11. Next, the methyl transferase genes gsmt and sdmt from Aphanothece halophytica were introduced into Bp1Z11 to construct the engineered strain Bp1Z12, which achieved a GB yield of 2.56 mg/L (a 326.7% increase compared with the wild type) after 36 h of fermentation. Under 0.3 mol/L NaCl stress (simulating moderately salinized soil conditions), the engineered strain achieved a GB yield of 4.94 mg/L (a 93.7% increase compared with the salt-free control) after 36 h of fermentation, with the biomass (OD600) increasing to 16.22 (a 20.9% increase compared with the salt-free control). Additionally, Bp1Z12 effectively alleviated salt stress of tomato plants and enhanced their growth in salinized soil. The root length of tomato plants in the Bp1Z12 treatment increased significantly by 75.0% and 27.3% compared with that in the water and Bp1 treatments, respectively. The plant height of the Bp1Z12 treatment increased by 76.9% and 21.1% compared with that in the water and Bp1 treatments, respectively. The leaf area of this treatment increased by 77.8% and 45.0% compared with that in the water and Bp1 treatments, respectively. The engineered strain Bp1Z12 can efficiently utilize glucose to synthesize GB, while exhibiting good salt tolerance and plant growth-promoting ability in salinized soils. This study provides new ideas for application of this strain in the development of stress-tolerant microbial fertilizers or the remediation of salinized soils in the future. 盐渍化土地给生态和农业造成了极大影响。生物修复是盐渍化地治理的重要绿色手段，提高根际促生菌(plant growth-promoting rhizobacteria, PGPR)耐盐性是提升生物修复效果的关键。甘氨酸甜菜碱(简称甜菜碱，glycine betaine, GB)作为重要的渗透压调节剂，可增强微生物和植物的抗逆性。为提高PGPR在盐渍化土壤中的存活率，本研究采用生物法合成GB，使用新型PGPR副地衣芽孢杆菌(Bacillus paralicheniformis) Bp1为底盘菌株，改造GB合成途径。首先，敲除甘氨酸竞争/消耗途径中的相关基因maeA、aceB、iclR和gcvP，强化乙醛酸循环(引入大肠杆菌来源基因aceAK)，引入外源高效转氨酶基因agx1，得到菌株Bp1Z09，使GB前体甘氨酸在发酵30 h内最高产量达到34.27 mg/L，较野生型12 h时的最高产量提升293.9%。同时，为保证菌株稳定性，将aceAK整合至基因组得到菌株Bp1Z11。之后，在Bp1Z11中引入嗜盐隐杆藻(Aphanothece halophytica)甲基化酶基因gsmt和sdmt，构建GB合成工程菌Bp1Z12，发酵至36 h时GB产量为2.56 mg/L，较野生型提高326.7%。在0.3 mol/L NaCl (模拟中高度盐渍化土壤环境)胁迫下，工程菌株GB产量在发酵至36 h时达4.94 mg/L (较无盐对照提升93.7%)，生物量(OD600)增至16.22 (较无盐对照提升20.9%)，凸显了其在盐渍化土壤微生物修复中的应用潜力。此外，Bp1Z12在盐渍化土中可有效缓解番茄受到的盐胁迫，提高番茄在盐渍化土中的生长能力，Bp1Z12处理组的根长较H2O和Bp1处理组分别显著增加75.0%和27.3%，株高较H2O和Bp1组分别增加76.9%和21.1%，叶面积较H2O和Bp1组分别增加77.8%和45.0%。工程菌Bp1Z12能高效利用葡萄糖合成GB，兼具良好耐盐性与促植物耐盐能力，为后续以该菌株为基础开发抗逆微生物肥料或土壤处理剂及进行盐渍化土壤修复提供了新的思路。.

The aim of this study was to characterize the molecular structure of the erb-b2 receptor tyrosine kinase 2 (ERBB2) gene and its expression in different parts of the oviducts of yaks in different phases of an oestrous cycle. We collected yak oviduct tissue samples, cloned the coding sequence (CDS) of ERBB2, and analyzed the physicochemical properties of ERBB2. The expression of ERBB2 in different parts of oviducts of yaks in different phases of an oestrous cycle were determined by RT-qPCR. The expression of ERBB2 protein in different parts of oviducts of yaks in different phases of an oestrous cycle was detected by Western blotting and immunohistochemistry. The results showed that the CDS of ERBB2 from yaks was 3 768 bp in size, encoding 1 255 amino acid residues. This gene was most closely related to that from yellow cows and most distantly related to that from camels. ERBB2 was a hydrophilic protein, with the molecular formula of C6464H10145N1815O1931S81 and the relative molecular mass of 146 kDa. Its secondary structure was dominated by random coils. The gene was differentially expressed in different parts of the oviducts in follicular and luteal phases, which indicated that it may be involved in reproductive processes such as sperm-egg binding and embryonic development. The findings provide a theoretical basis for further research on the functions and molecular mechanisms of ERBB2 in female yak reproduction. 本研究旨在探究表皮生长因子受体2 (erb-b2 receptor tyrosine kinase 2, ERBB2)基因的分子结构特征及其在牦牛不同发情周期输卵管不同部位中的表达情况。采集牦牛输卵管组织样品，对牦牛ERBB2基因编码区序列(coding sequence, CDS)进行克隆，分析ERBB2的理化性质等;应用实时荧光定量PCR (real-time fluorescence quantitative PCR, RT-qPCR)检测ERBB2基因在牦牛发情周期不同时期输卵管各段中的表达情况;应用蛋白免疫印迹法(Western blotting, WB)和免疫组织化学法(immunohistochemistry, IHC)检测发情周期不同时期输卵管各部位中ERBB2蛋白的表达。结果显示，牦牛ERBB2基因编码区片段大小为3 768 bp，共编码1 255个氨基酸，与黄牛的亲缘关系最近，与骆驼亲缘关系最远。牦牛ERBB2蛋白属于亲水性蛋白，ERBB2基因编码蛋白分子式为C6464H10145N1815O1931S81，相对分子质量146 kDa，其二级结构以不规则卷曲为主。该基因在卵泡期和黄体期这2个时期的输卵管各段中的表达存在差异，可能参与精卵结合以及胚胎发育等生殖过程。本研究为进一步研究ERBB2在雌牦牛繁殖中的功能和分子机制提供了理论依据。.

Chiral amines as pivotal intermediates in organic synthesis are widely utilized in the production of pharmaceuticals, fine chemicals, and bioactive molecules, holding significant industrial value. The asymmetric synthesis of chiral amines has always gained great attention. NAD(P)H-dependent imine reductases (IREDs) with wide substrate ranges, high activity, and high enantiomeric selectivity can be used for asymmetric reduction of imines to chiral amines. Our study aims to develop an efficient and stable immobilized dual-enzyme system through the fusion expression of imine reductase and formate dehydrogenase, in order to address the challenge of coenzyme regeneration and enhance catalytic efficiency, thereby providing a novel strategy for the green synthesis of chiral amines. The constructed system was applied to catalyze the asymmetric reduction of 1-methyl-3,4-dihydroisoquinoline to synthesize (S)-1-methyl-1,2,3,4-tetrahydroisoquinoline. Comparative analysis indicated that the catalytic efficiency of this fusion expression system exceeded that of both co-expression and standalone dual-enzyme systems. Furthermore, mesoporous silica nanoflowers were utilized as carriers to immobilize the fusion-expressed dual enzymes through a covalent method. In the case of covalent binding duration of 1.5 h and an initial enzyme concentration of 2.5 mg/mL, the protein loading achieved 193.2 mg/g. The immobilized enzymes demonstrated excellent pH, thermal, and storage stability. When the immobilized enzymes were employed to catalyze asymmetric reduction reactions of other cyclic imines, such as 1-ethyl-3,4-dihydroisoquinoline, 5-phenyl-3,4-dihydro-2H-pyrrole, 2,3,3-trimethyl-3H-indole, 2,3,3,5-tetramethylindole, and myosmine, the conversion rates exceeded 95%, and the values of e.e. surpassed 96%. The data confirm the application potential of the immobilized fusion enzymes in the green and efficient synthesis of chiral amines. Our study provides a novel strategy for the industrial biosynthesis of chiral amines, and the developed fused-enzyme immobilization approach holds significant theoretical and practical value for addressing common technical challenges in cofactor-dependent biocatalytic processes. 手性胺作为关键的有机合成中间体，广泛应用于药物、精细化学品及生物活性分子的合成，具有重要的工业价值。手性胺的不对称合成一直备受关注，NAD(P)H依赖型亚胺还原酶(imine reductase, IRED)具有底物范围广、高活性和立体选择性的特点，可用于不对称还原合成手性胺。本研究旨在通过构建一种高效、稳定的IRED与甲酸脱氢酶(formate dehydrogenase, FDH)双酶融合表达与固定化体系，解决辅酶再生难题并提升催化效率，为手性胺的绿色合成提供新策略。该体系用于催化1-甲基-3,4-二氢异喹啉不对称还原合成(S)-1-甲基-1,2,3,4-四氢异喹啉。通过对比发现，融合表达体系的催化效果优于共表达和单独表达双酶的体系。其次，利用介孔氧化硅纳米花(mesoporous silica nanoflowers, MSN)作为载体，共价法固定化融合表达双酶，固定化时间为1.5 h且初始酶浓度为2.5 mg/mL时，蛋白负载量为193.2 mg/g。固定化酶展现出良好的pH稳定性、热稳定性和储藏稳定性。将制备的固定化酶应用于催化其他环状亚胺如1-乙基-3,4-二氢异喹啉、5-苯基-3,4-二氢-2H-吡咯、2,3,3-三甲基-3H-吲哚、2,3,3,5-四甲基吲哚和麦斯明等的不对称还原反应，转化率高于95%，对映体过量值(enantiomeric excess, e.e.)高于96%，验证了固定化融合酶催化体系在手性胺绿色高效合成中的应用潜力。本研究为手性胺的工业化生物合成提供了新思路，所开发的融合酶固定化策略对解决辅酶依赖型生物催化过程中的共性技术难题具有重要的理论与应用价值。.

The carboxyl-terminal domain of the equine herpesvirus-1 (EHV-1) gM gene has been shown to induce the production of gM vesicles capable of loading or displaying target proteins in mammol/Lalian cells. These vesicles show promising potential for applications in targeted drug delivery, gene therapy, and the development of enveloped virus-like particle (eVLP) vaccines. However, the biogenesis, structural composition, and biological functions of gM vesicles remain poorly understood. To address this, we employed immol/Lunoaffinity capture to purify gM vesicles and characterized their morphology, concentration, and zeta potential by transmission electron microscopy (TEM) and nanoparticle tracking analysis (NTA). In parallel, the protein composition of gM vesicles was identified by liquid chromatography-tandem mass spectrometry (LC-MS/MS), followed by functional annotation via GO (gene ontology) and pathway analysis using KEGG (Kyoto encyclopedia of genes and genomes). The results demonstrated that gM vesicles exhibited a diameter range of 100‒300 nm, an average diameter of (151.3±5.7) nm, a zeta potential of ‒32.579 mV, and a yield of approximately 3.85×105 particles/mL. A total of 416 proteins were identified by LC-MS/MS. Following the exclusion of unannotated proteins, 396 proteins were remained for subsequent GO and KEGG analyses. The results revealed that gM vesicles were involved in four metabolic pathways associated with vesicle formation and trafficking: vesicle-mediated transport, endocytosis, endoplasmic reticulum-Golgi transport, and protein processing within the endoplasmic reticulum. This study elucidates the formation mechanism of gM vesicles at the protein level, thereby establishing a foundation for their future applications in various fields, such as modification of gM vesicles, regulation of intracellular vesicle dynamics, drug delivery, and targeted therapy. 马疱疹病毒-1型(equine herpesvirus-1, EHV-1) gM基因的羧基端可诱导哺乳动物细胞产生能够负载或展示靶蛋白质的囊泡(gM囊泡)，有望应用于药物靶向递送、基因治疗及囊膜化病毒样颗粒疫苗研发等领域。为了研究该囊泡的形成机制、结构构成及生物学功能，本研究使用免疫亲和技术纯化gM囊泡，通过透射电子显微镜(transmission electron microscope, TEM)、纳米颗粒跟踪分析技术(nanoparticle tracking analysis, NTA)对纯化的gM囊泡的形态、数量及Zeta电位进行分析;同时，利用蛋白质质谱技术(liquid chromatography-tandem mass spectrometry, LC-MS/MS)鉴定gM囊泡的结构蛋白，并对蛋白质鉴定结果进行基因本体(gene ontology, GO)功能注释和KEGG (Kyoto encyclopedia of genes and genomes)代谢通路分析。结果显示，gM囊泡直径分布在100‒300 nm之间，其平均直径约(151.3±5.7) nm;囊泡Zeta电位为‒32.579 mV;纯化囊泡总量可达3.85×105 particles/mL细胞;利用蛋白质质谱技术共鉴定出416种蛋白质，去除无注释的蛋白后保留396种蛋白质用于后续分析;通过GO及KEGG分析，gM囊泡共参与了4条与囊泡形成及运输过程相关的代谢通路，分别为囊泡介导的运输、内吞作用、内质网与高尔基体间的运输和内质网中的蛋白质加工。本研究通过蛋白质谱鉴定分析从蛋白质水平上揭示了gM囊泡的形成机制，为gM囊泡在工程化改造、胞内调控机制、药物递送靶向治疗等领域的研究及应用奠定基础。.

This study aims to establish a double-antigen sandwich ELISA method for detecting serum antibodies against caprine parainfluenza virus type 3(CPIV3), thereby supporting the clinical diagnosis of CPIV3 infection. According to the reference nucleocapsid (N) gene sequence (NCBI accession number: MT756864.1) published in NCBI, the N gene was synthesized and inserted into pET-30a to construct the recombinant plasmid pET-30a-N. The N protein was expressed in Escherichia coli and purified by Ni2+-affinity chromatography via its His tag. Reactivity of the purified N protein was verified by indirect ELISA and Western blotting. The His tag was removed with thrombin (1:1 000, W/W). The optimal coating concentration of the capture antigen, dilution of the HRP-conjugated detection antigen, and serum dilution were determined by checkerboard titration. Ultimately, a double-antigen sandwich ELISA method was established for clinical CPIV3 samples. Indirect ELISA and Western blotting confirmed that the N protein exhibited excellent antigenicity. The His tag was completely removed without altering the reactivity of the N protein. Checkerboard assays established the following optimal parameters: coating antigen concentration of 0.5 µg/mL, HRP-conjugated antigen dilution of 1:10 000, and serum dilution of 1:20. The cut-off value for the assay was set at 0.237 (OD450). No cross-reactivity was observed with positive sera against foot-and-mouth disease virus type O (FMDV-O), sheep/goat pox virus (GTPV), orf virus (ORFV), or Brucella. The sensitivity of the developed ELISA reached 1:512, being significantly higher than that of a commercial kit. Both intra- and inter-assay coefficients of variation were below 10%. Parallel testing of 150 field sheep sera with the Shengbo indirect ELISA kit yielded an overall concordance rate of 87%. The established double-antigen sandwich ELISA method demonstrates high specificity, sensitivity, and reproducibility. It provides an effective technical tool for large-scale surveillance, sero-epidemiological investigations, and disease control of CPIV3 infection. 本研究旨在建立一种双抗原夹心ELISA方法，用以检测羊副流感病毒3型血清抗体，以支持羊副流感病毒3型血清抗体感染临床诊断。合成N基因(NCBI数据库登录号:MT756864.1)并构建至pET-30a中，命名为pET-30a-N，利用大肠杆菌原核表达、亲和层析纯化N蛋白。用间接ELISA方法和Western blotting方法验证N蛋白反应性。用凝血酶(1:1 000，W/W)酶切N蛋白His标签。通过优化抗原包被浓度、酶标抗原稀释比例、血清稀释比例等条件，最终建立用于检测临床样本羊副流感病毒3型血清抗体的双抗原夹心ELISA方法。结果发现，间接ELISA方法和Western blotting方法鉴定N蛋白具有良好的反应性。N蛋白His标签成功切除，且酶切后的N蛋白反应性未改变。通过棋盘法得到最佳抗原包被浓度为0.5 μg/mL;HRP酶标抗原稀释度为1:10 000;血清最佳稀释比例1:20;双抗原夹心ELISA临界值为0.237;与口蹄疫病毒O型(foot-and-mouth disease virus, FMDV-O)、羊痘病毒(sheep/goat pox virus, GTPV)、羊口疮病毒(orf virus, ORFV)、布鲁氏菌(Brucella)的抗体阳性血清均无交叉反应，与商品化试剂盒相比较敏感性高达1:512;批间和批内的变异系数均小于10%。用本研究建立的方法与圣博羊间接ELISA试剂盒同时对150份田间羊临床血清进行检测，两者的符合率为87%。本研究所建立的双抗原夹心ELISA方法具备特异性强、敏感性高、重复性好等特点，为羊副流感病毒3型血清抗体大规模样品监测、血清学调查、疫病防控提供了有效的技术手段。.

Duck enteritis virus (DEV) is a common herpesvirus that can cause acute, febrile, and septicemic infectious diseases in poultry, leading to tremendous economic losses in the poultry farming industry. Therefore, there is an urgent need for efficient and sensitive on-site rapid detection of DEV. Targeting the conserved fragment of the UL2 gene of DEV, specific recombinase polymerase amplification (RPA) primers and probes were designed. After optimization of the RPA reaction conditions, a fluorescent RPA isothermal rapid detection method for DEV was established. The results demonstrated that this detection method had the optimal reaction temperature of 40 ℃ and the optimal reaction time of 20 min, with a minimum detection limit of 2 fg/reaction for the virulent strain of DEV. The method specifically amplified only the virulent strain of DEV, showing no cross-reactivity with other common duck pathogens such as duck hepatitis virus and duck Tembusu virus. The established real-time fluorescent RPA method and qPCR were used to test 36 clinical samples of duck tissue. The RPA method demonstrated the sensitivity of 100%, specificity of 100%, and accuracy of 100%. The real-time fluorescent RPA detection method established in this study for the virulent strain of DEV features rapid detection, high sensitivity, and strong specificity, providing a feasible approach for the on-site rapid detection of duck plague. 鸭瘟病毒(duck enteritis virus, DEV)是一种常见的疱疹科病毒，可引起禽类急性、热性、败血性传染病，造成家禽养殖业巨大的经济损失。因此，迫切需要对鸭瘟病毒进行高效、灵敏的现场快速检测。以DEV的UL2基因保守片段为靶基因，设计特异性重组酶聚合酶扩增(recombinase polymerase amplification, RPA)引物和探针，优化RPA反应条件后，建立DEV的荧光RPA恒温快速检测方法。结果证明，该检测方法最佳反应温度为40 ℃，最佳反应时间为20 min，对鸭瘟强毒株的最低检测限为2 fg/反应;仅特异性扩增DEV强毒株，与其他常见鸭病病原如鸭肝炎病毒、鸭坦布苏病毒等均无交叉反应。采用建立的实时荧光RPA检测方法和实时荧光定量聚合酶链式反应(quantitative real-time polymerase chain reaction, qPCR)对36份鸭组织临床样本进行检测，结果显示RPA检测敏感性为100%，特异性为100%，准确率为100%。本研究建立的DEV强毒实时荧光RPA检测方法，具有检测速度快、灵敏度高、特异性强等特点，为鸭瘟疫病的现场快速检测提供了一种可行的途径。.

Pectinases play a crucial role in the enzymatic degumming process of ramie. However, existing pectinases often suffer from issues such as low degumming efficiency and long processing time. Therefore, it is essential to discover novel efficient pectinases for the ramie degumming process. In this study, we performed carbohydrate-active enzymes annotation on the coding sequence (CDS) and protein sequence of Bacillus licheniformis ATCC 14580, identifying a pectate lyase BlicPL1 whose function has not yet been verified. An expression vector for this gene was constructed and expressed in Escherichia coli BL21(DE3), followed by protein purification. This study marks the first experimental confirmation that BlicPL1 functions as a pectate lyase, with a specific enzyme activity of 3 952.54 U/mg, which is higher than those of most reported pectate lyases from the polysaccharide lyase 1 (PL1) family. Enzymatic characterization revealed that the optimal pH and temperature for BlicPL1 activity were 10.0 and 60 ℃, respectively. Notably, BlicPL1 maintained high activity (over 80% relative activity) within a broad range of 50-80 °C and pH 10.0-12.0. This thermostable and alkali-tolerant profile aligns well with the high-temperature, high-pH conditions typical of ramie degumming processes. Unlike the conventional pectate lyases which are activated by Ca2+, BlicPL1 showcased the activity not promoted by Ca2+, or even inhibited by Ca2+, which highlighted its uniqueness in the Ca2+ response mechanism and suggested that BlicPL1 was a novel pectate lyase. Furthermore, BlicPL1 was employed for ramie degumming, resulting in a weight loss rate of 16.17% and a degumming rate of 44.09% for ramie fibers. Scanning electron microscopy results indicated that the treatment with BlicPL1 helped remove residual middle lamella tissue from the ramie fiber surface and reduced the adhesion of free gummy substances, resulting in smoother and more separated fibers. X-ray diffraction analysis confirmed that the characteristic cellulose peaks of the ramie fibers remained unchanged after degumming by BlicPL1, while the crystallinity index increased from 74.59% to 79.69%, indicating an increased proportion of cellulose and the successful removal of non-cellulosic components. The pectate lyase BlicPL1 identified in this study, with high activity and suitability for medium-high temperatures and alkaline environments, demonstrates significant application potential in the ramie degumming process. This work lays a foundation for advancing the industrial application of pectinases in ramie degumming. 果胶酶在苎麻酶法脱胶过程中发挥重要作用，但现有果胶酶存在脱胶效率低、脱胶时间长等问题。为了挖掘新型、高效的果胶酶应用于苎麻脱胶工艺，本研究对地衣芽孢杆菌ATCC 14580的编码序列(coding sequence, CDS)和蛋白序列进行碳水化合物酶注释，发掘了一个尚未验证功能的果胶裂解酶BlicPL1。构建BlicPL1基因的大肠杆菌BL21(DE3)表达载体，并进行了诱导表达和分离纯化。本研究首次通过实验证实了BlicPL1的果胶裂解酶活性，其比酶活为3 952.54 U/mg，高于多数已报道的多糖裂解酶1 (polysaccharide lyase 1, PL1)家族果胶裂解酶。酶学性质研究显示果胶裂解酶BlicPL1最适反应pH为10.0，最适反应温度为60 °C;且在反应温度50-80 °C和pH 10.0-12.0范围内都表现出较高的酶活性，相对酶活力保持在80%以上。果胶裂解酶BlicPL1耐高温、耐碱特性与苎麻脱胶工艺中的高温、高pH环境具有很好的适配性。与常规果胶裂解酶依赖Ca2+激活不同，Ca2+对果胶裂解酶BlicPL1的活性不仅无明显促进作用，甚至有一定抑制作用，凸显了其在Ca2+响应机制方面的独特性，表明果胶裂解酶BlicPL1是一种新型果胶裂解酶。进一步将果胶裂解酶BlicPL1用于苎麻脱胶，苎麻失重率为16.17%，苎麻脱胶率达到44.09%;扫描电子显微镜结果显示果胶裂解酶BlicPL1作用后有助于去除苎麻表面残留的中间层状组织并减少游离胶状物质附着，使苎麻表面变得更光滑、纤维变得更加分散;X射线衍射结果显示果胶裂解酶BlicPL1脱胶后苎麻纤维特征纤维素峰位不变，结晶度从74.59%升至79.69%，表明纤维素占比增加，非纤维素物质被去除。本研究发掘的果胶裂解酶BlicPL1具有活性高，适用于中高温、碱性环境等特点，在苎麻脱胶工艺中具有较大应用潜力，为加速推进果胶酶运用于工业苎麻脱胶工艺奠定了基础。.

Phage therapy has the potential for broad application in the treatment of mastitis in dairy cows and the control of drug-resistant bacterial infections because of its precise targeting, efficient lytic activity, self-replicating nature, and excellent biocompatibility. This review summarized the challenges and future prospects in the utilization of phage therapy for bovine mastitis. The review encompassed the isolation and identification of phages targeting major mastitis pathogens (including Staphylococcus aureus and Escherichia coli) and the evaluation of in vivo (animal models and dairy cows) and in vitro (bacteriostatic effects, biofilm clearance, etc.) effectiveness. The findings indicate that specific phages or cocktail preparations are effective in controlling target pathogens. Phage genomic analysis is instrumental in elucidating the cleavage mechanism and evaluating potential virulence/lysogenic genes, thereby ensuring the safety of the treatment. Concurrently, the review delves into the prevailing challenges in this domain, encompassing the limited spectrum of phage hosts, the intricacies of pharmacokinetics and optimization of drug delivery in vivo, substantial-scale manufacturing concerns, and stability considerations. Notwithstanding the numerous challenges, innovative directions, such as phage engineering, phage-antibiotic synergistic application, and new delivery systems (such as nanocarriers), provide important ideas for future development. This review aims to offer theoretical foundations and practical references for further research and application of phage therapy in the prevention and control of bovine mastitis, promoting the translation of this green treatment strategy into clinical practice. 噬菌体疗法因精准靶向性、高效裂解性、自我增殖性及良好生物相容性，在奶牛乳房炎治疗和耐药菌感染控制领域具有广阔的应用前景。本文综述了在利用噬菌体治疗奶牛乳房炎过程中所面临的挑战与未来前景，总结了主要乳房炎病原体(金黄色葡萄球菌、大肠杆菌等)噬菌体的分离鉴定方法以及已开展的体内(实验动物模型及奶牛)和体外(抑菌效果、细菌生物膜清除等)有效性评价研究，归纳了特定噬菌体或鸡尾酒制剂在控制目标病原体方面的效果，阐述了噬菌体基因组学分析在理解裂解机制、评估潜在毒力/溶原性基因中起的关键作用。同时，深入探讨了该领域当前面临的挑战，包括噬菌体宿主范围相对狭窄、体内药代动力学及给药方案优化困难、规模化生产与稳定性问题等。尽管挑战众多，但是噬菌体工程改造、噬菌体-抗生素协同应用、新型递送系统(如纳米载体)等创新方向仍为未来发展提供了重要研究思路。本文为噬菌体疗法在奶牛乳房炎防控中的进一步研究与应用提供了理论依据和实践参考，推动该绿色治疗策略向临床转化。.

We had engineered Caldicellulosiruptor morganii CE (CmCE) to construct the enzyme CmCE/D226G for catalyzing the isomerization of lactose to lactulose. However, the poor thermostability and catalytic efficiency of CmCE/D226G limits its application under high substrate concentrations. To address these issues, we employed directed evolution to improve the catalysis performance of CmCE/D226G. First, we used error-prone PCR to construct a variant library and identified the positive variants through cysteine-tryptophan chromogenic high-throughput screening. Then, aiming the crucial residue positions, we conducted saturation mutagenesis to obtain the variants with improved thermostability and catalysis. Accordingly, we successfully obtained a variant named CmCE/D226G/S180T. Its half-life and catalysis efficiency (kcat/Km) at 70 ℃ were 184.2 min and 36.5 L/(mmol·s), respectively, which were higher than those (117.7 min and 14.96 L/(mmol·s), respectively) of CmCE/D226G. The improvement of isomerization efficiency resulted from the shortening distance between His377 and the O1 of substrate enol intermediate. The improvement of thermostability was attributed to the rigidness enhancement of a flexible loop. With 350 g/L lactose as the substrate, CmCE/D226G/S180T achieved a lactulose yield of 54.8% and an epilactose yield of 9.2%, while CmCE/D226G only reached a lactulose yield of 50% and an epilactose yield of 10.3%. Taken together, this work provides an elite enzyme for catalyzing the transformation of lactose to lactulose, which especially facilitates the innovation of eco-friendly synthesis of lactulose. 来自热解纤维素菌(Caldicellulosiruptor morganii)的纤维二糖差向异构酶(CmCE)的突变体CmCE/D226G可以催化乳糖异构化为乳果糖，但热稳定性和催化活性制约了其在高底物浓度下的生物转化。为解决上述问题，本研究针对CmCE/D226G开展定向进化，采用易错PCR建立突变体文库，结合盐酸半胱氨酸-色氨酸显色高通量筛选正向突变体，再针对关键位点开展饱和突变，获得了热稳定性和催化活性提高的突变体CmCE/D226G/S180T。该酶在70 ℃下的半衰期从117.7 min提高到184.2 min，催化效率从14.96 L/(mmol·s)提高到36.5 L/(mmol·s)。机理分析表明，突变酶的His377和底物烯醇式中间体O1的距离更近，从而提升了异构化效率，同时突变增强了柔性环刚性，提高了酶热稳定性。以350 g/L乳糖进行生物转化，CmCE/D226G/S180T的乳果糖得率达54.8%，依匹乳糖得率为9.2%;相比之下，CmCE/D226G的乳果糖和依匹乳糖得率仅分别为50%和10.3%。本研究为生物法制备乳果糖提供了优势酶，有利于促进乳果糖的绿色合成技术创新。.

Selenium is an essential trace element for the human body, enhancing the body resistance to a variety of diseases induced by oxidative stress. Selenium nanoparticles (SeNPs) possess advantages such as low toxicity and broad biocompatibility. In order to screen for a strain with high efficiency in synthesizing SeNPs, we investigated the selenium tolerance and SeNPs synthesis capacities of three selenium-tolerant yeast strains and the in vitro antioxidant activities and antimicrobial activities of the synthesized SeNPs. Furthermore, we characterized SeNPs synthesized by strain M16-28 with the best performance. The results indicated that strain M16-28 was capable of tolerating 300 mmol/L Na2SeO3, with a SeNPs synthesis capacity of 1 018.92 μg/mL, which was 2.86 times that of strain M1. The in vitro antioxidant experiments indicated that the synthesized SeNPs exhibited the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical scavenging rate of 65.7%, the hydroxyl radical scavenging rate of 57.5%, and the reducing power of 0.77. The antimicrobial experiments indicated that the synthesized SeNPs displayed significantly stronger inhibitory effects against Gram-positive bacteria than against Gram-negative bacteria, and they had a significant inhibitory effect on the pathogenic microorganisms causing stem rot of Lycium barbarum. The SeNPs synthesized by the three yeast strains exhibited an inhibition rate higher than 90% against Fusarium spp., and a lower inhibition rate against Alternaria alternata.The SeNPs produced by strain M16-28 exhibited the highest antimicrobial activity, with an inhibition rate of (66.53±1.37)%. Characterization results indicated that SeNPs were closely arranged, encapsulated by polysaccharides, and negatively charged, with a size ranging from 190 nm to 390 nm and the zeta potential of -7.87 mV. In conclusion, strain M16-28 demonstrates ideal performance of synthesizing SeNPs, and the capacity for selenium enrichment is positively correlated with the ability to synthesize SeNPs. This study provides a potential microbial resource for the efficient synthesis of SeNPs. 硒是人体必需的微量元素，对许多由氧化应激引起的疾病具有抗性。纳米硒(selenium nanoparticles, SeNPs)具有低毒和广泛生物活性的优点。为了筛选高效合成SeNPs的菌株，本研究对3株耐硒酵母菌的硒耐受性、SeNPs合成能力、体外抗氧化活性和抑菌活性进行研究，选择出性能最好的菌株M16-28进行表征。结果表明，M16-28菌株能耐受300 mmol/L Na2SeO3，其合成SeNPs的能力达1 018.92 μg/mL，是M1菌株合成SeNPs含量的2.86倍。体外抗氧化实验表明，1,1-二苯基-2-三硝基苯肼(1,1-diphenyl-2-picrylhydrazyl DPPH)自由基清除率为65.7%，羟自由基清除率为57.5%，还原力为0.77。抑菌性试验表明，对革兰氏阳性菌的抑制效果显著高于革兰氏阴性菌，对枸杞茎腐病病原微生物具有显著抑制作用。3株菌合成的SeNPs对镰刀菌属的抑制率均高于90%，对链格孢菌株抑制率较低，M16-28菌株合成的SeNPs抑菌性最高，为(66.53±1.37)%。表征结果表明，SeNPs呈紧密排布，有多糖包裹，带负电荷，尺寸分布在190-390 nm之间，Zeta电位为-7.87 mV。综上所述，M16-28菌株具有较好的合成SeNPs能力，富硒能力和合成SeNPs能力呈正相关，可作为高效合成SeNPs的潜在微生物资源。.

Hyaluronic acid (HA) is a glycosaminoglycan with significant biological activities, which render it widely applicable in the cosmetics and pharmaceutical industries. The development of safe and efficient chassis cells to enhance HA synthesis efficiency has thus emerged as a key factor in HA production. Our study aims to construct a high-performance HA biosynthesis system using Streptomyces as the chassis cell, thereby providing technical support for the efficient microbial production of HA. Thus, our study focused on the metabolic engineering of Streptomyces for strengthening the HA synthesis pathway and then optimized the culture conditions for efficient HA synthesis. First, the HA-synthesizing capabilities of four hyaluronate synthases from different sources were evaluated in two host strains: Streptomyces coelicolor M1146 and Streptomyces albus J1074. The results indicated that the hyaluronate synthases derived from Streptococcus pyogenes exhibited the strongest HA synthesis capability. Notably, the HA yield in S. albus J1074 was higher. Building on this finding, S. albus J1074 was selected as the starting strain to construct a chassis strain tailored for HA synthesis: key genes in the competitive metabolic pathway of HA synthesis were knocked out, while the expression levels of genes associated with the bypass pathway were down-regulated. Furthermore, different combinations of key genes involved in the HA precursor synthesis pathway were designed, and their expression levels were enhanced via a constitutive strong promoter. The recombinant strain obtained therefrom achieved a maximum HA yield of 2.62 g/L. Finally, the synthetic capacity of this high-yield engineered strain was further unleased through the optimization of culture conditions, leading to a final HA yield of 4.63 g/L. The recombinant strain constructed in this study not only lays a foundation for the development of engineered Streptomyces but also provides an excellent chassis strain for the microbial production of HA and other related bioproducts. 透明质酸是一种具有重要生物活性的糖胺聚糖，在化妆品和医药领域具有广泛应用。开发安全高效的底盘细胞以提升透明质酸的合成效率，已成为透明质酸生产的关键。本研究旨在构建以链霉菌为底盘细胞的高性能透明质酸合成体系，为透明质酸的微生物高效合成提供技术支撑。本研究主要聚焦链霉菌的代谢工程改造，通过强化透明质酸合成通路，结合培养条件的优化，最终实现透明质酸的高效合成。首先，在天蓝色链霉菌和小白链霉菌这2种宿主中，分别测试了4种不同来源的透明质酸合成酶的合成能力。结果表明:在这2种链霉菌中，源自化脓性链球菌的透明质酸合成酶的合成能力最强，且在小白链霉菌J1074中产量更高。随后，以小白链霉菌J1074为出发菌株，通过敲除透明质酸合成竞争代谢途径的关键基因，同时下调旁路途径相关基因的表达水平，构建了适用于透明质酸合成的底盘细胞。此外，对透明质酸前体合成途径中的关键基因进行了不同组合，并利用组成型强启动子强化其表达水平，获得的重组菌株的透明质酸产量最高达到2.62 g/L。最后，通过优化培养条件进一步释放工程菌株的合成能力，最终透明质酸产量提升至4.63 g/L。本研究构建的重组菌株，不仅为工程链霉菌的开发奠定了基础，也为透明质酸及类似生物产品的发酵制备提供了优异的底盘菌株。.

This study aimed to explore the regulatory effect and mechanism of 3β-hydroxy-Δ5-C27-steroid oxidoreductase (HSD3B7) on bovine follicular development. Bovine primary granulosa cells (GCs) with HSD3B7 knockdown and overexpression were constructed, and the proliferation efficiency of GCs was examined by the cell counting kit-8 (CCK-8) method. Annexin V-EGFP/PI double staining was performed to detect the apoptosis level of GCs. The concentration of estradiol (E2) in the culture medium and that of 2-methoxyestrone in GCs were determined by enzyme-linked immunosorbent assay (ELISA). The relative expression levels of the CCND2, Bim, Caspase-3, CYP19A1 and STAR were determined by qRT-PCR. The protein levels of related genes were determined by Western blotting. The results showed that HSD3B7 knockdown decreased the proliferation efficiency and CCND2 expression level of primary bovine follicular GCs, promoted the apoptosis, and increased Bim and Caspase-3 expression in GCs. Additionally, it decreased E2 secretion and CYP19A1 and STAR expression, and increased the concentration of 2-methoxyestrone, the main metabolite of E2. The opposite results were obtained after HSD3B7 overexpression. These results suggested that HSD3B7 positively regulated the development of bovine follicles by promoting the proliferation and E2 secretion and inhibiting the apoptosis of GCs. The results provide a new theoretical basis for further understanding the molecular regulatory network of bovine follicle development, and also lay a foundation for related research and practical application to improve bovine reproductive efficiency. 本研究旨在探究3β-羟基-Δ5-C27-类固醇氧化还原酶(3β-hydroxy-Δ5-C27-steroid oxidoreductase, HSD3B7)对牛卵泡发育的调节作用及其机制。构建敲降和过表达HSD3B7基因的牛原代颗粒细胞(granulosa cells, GCs)，用细胞计数试剂-8 (cell counting kit-8, CCK-8)法检测GCs增殖效率，用Annexin V-EGFP/PI双染法检测GCs凋亡水平，用酶联免疫吸附试验(enzyme-linked immunosorbent assay, ELISA)技术检测培养液中雌二醇(estradiol, E2)和胞内2-甲氧基雌酮的浓度，用实时荧光定量PCR (real-time quantitative-polymerase chain reaction, qRT-PCR)技术检测CCND2、Bim、Caspase-3、CYP19A1、STAR的相对表达水平，用Western blotting技术检测相关基因蛋白水平的表达情况。结果显示，HSD3B7敲降后降低了牛卵泡原代GCs增殖效率和CCND2蛋白的表达水平;促进了GCs凋亡和Bim蛋白、Caspase-3蛋白的表达;减少E2的分泌量和CYP19A1蛋白、STAR蛋白的表达量;增加E2主要代谢物2-甲氧基雌酮的浓度。HSD3B7过表达后取得相反结果。本研究结果表明，HSD3B7通过促进牛GCs增殖和E2分泌、抑制GCs凋亡，正向调控牛卵泡发育进程。研究结果为深入解析牛卵泡发育的分子调控网络提供了新的理论依据，也为提高牛繁殖效率的相关研究和实践应用奠定了基础。.

In the special marine environment characterized by high salinity, high osmotic pressure, and oligotrophy, marine actinomycetes have evolved unique survival strategies. Some of their secondary metabolites, with novel structures and potent biological activities, hold significant research and application value in pharmaceutical development. Under conventional laboratory culture conditions, only 20% of the biosynthetic gene clusters (BGCs) in marine actinomycetes are expressed, and a large number of silent BGCs urgently need to be explored. This paper systematically reviews the silencing mechanisms and activation strategies of silent BGCs in marine actinomycetes, comprehensively summarizes recent research advances in physical activation, chemical activation, and bioengineering technologies, and deeply analyzes the advantages and limitations of various activation methods. The silencing of BGCs often resulted from the combined effects of multiple factors. Therefore, this paper proposes a multi-technology combination strategy for activating silent BGCs. This strategy simulates the original survival environment of microorganisms from multiple aspects or constructs specific expression environments for gene clusters to promote and induce the expression of silent BGCs at multiple levels, thereby obtaining novel secondary metabolites. This strategy provides theoretical references and technical pathways for exploring the metabolic potential of marine actinomycetes and promoting the discovery of new natural products. 在海洋高盐、高渗、寡营养的特殊环境之中，海洋放线菌进化出了独特的生存策略，其部分次级代谢产物凭借新颖的结构和良好的生物活性，在医药开发领域具有重要的研究与应用价值。在实验室常规培养条件下，海洋放线菌仅有20%的生物合成基因簇(biosynthetic gene clusters, BGCs) 得以表达，大量处于沉默状态的BGCs亟待开发。本文系统梳理了海洋放线菌沉默BGCs的沉默机制与激活策略，全面总结近几年物理激活、化学激活、生物工程技术领域的研究成果，深入剖析各类激活方法的优势与局限性，并针对沉默BGCs的激活提出了多技术联用策略。多技术联用策略可从多方面模拟微生物原始生存环境或针对基因簇构造特定表达环境，多层面促进、诱导沉默BGCs表达，进而获得新颖次级代谢产物，为挖掘海洋放线菌代谢潜力、推动新型天然产物发现提供理论参考与技术路径。.

This study investigated the proteomic characteristics and biological functions of hypoxic exosomes derived from hypoxia-preconditioned feline adipose-derived mesenchymal stem cells (ADMSCs), aiming to reveal the remodeling effect of hypoxic preconditioning on the protein composition of exosomes derived from feline ADMSCs and its potential applications. CoCl2 was used to mimic a hypoxic environment for ADMSCs, and exosomes were isolated by differential ultracentrifugation. The physical properties of the exosomes were characterized by transmission electron microscopy, nanoparticle tracking analysis, and Western blotting. Proteomic analysis revealed that hypoxic preconditioning significantly altered the exosomal proteomic profile, identifying 120 differentially expressed proteins (116 upregulated and 4 downregulated). Bioinformatic analysis indicated significant enrichment of key pathways including the hypoxia-inducible factor-1 (HIF-1) signaling pathway, adenosine monophosphate-activated protein kinase (AMPK) signaling pathway, autophagy, and proteasome function. Gene ontology (GO) functional annotation demonstrated significant enrichment of biological processes such as metabolic processes, cell cycle regulation, and signal transduction in the hypoxia-preconditioned group. Kyoto encyclopedia of genes and genomes (KEGG) analysis further suggested potential biological functions through the regulation of pathways including the cell cycle and renin-angiotensin system. Notably, hypoxia-responsive proteins such as HMOX1 and TFRC were upregulated, while pathways related to the renin-angiotensin system were suppressed. This study systematically elucidates, for the first time, the remodeling effect of hypoxic preconditioning on the proteome of exosomes derived from feline ADMSCs, providing new molecular insights into exosome-mediated intercellular communication. 本研究旨在揭示低氧预处理对猫脂肪间充质干细胞(adipose-derived mesenchymal stem cells, ADMSCs)外泌体蛋白质组成的重塑作用及潜在应用。探讨了低氧预处理猫ADMSCs来源外泌体的蛋白质组特征及其生物学功能，通过CoCl{L-End} 2模拟低氧环境处理ADMSCs，采用差速超速离心法分离外泌体，经透射电镜、纳米颗粒追踪分析和蛋白质印迹鉴定其物理特性。蛋白质组学分析显示，低氧预处理诱导外泌体蛋白质组发生显著改变，共鉴定出120个差异表达蛋白质(116个上调、4个下调)。生物信息学分析表明，差异蛋白质显著富集于低氧诱导因子-1 (hypoxia-inducible factor-1, HIF-1)信号通路、腺苷酸活化蛋白激酶(adenosine monophosphate-activated protein kinase, AMPK)信号通路、自噬和蛋白酶体功能等关键通路。基因本体(gene ontology, GO)功能注释显示，低氧预处理组代谢过程、细胞周期调控和信号转导等生物学过程显著富集。京都基因与基因组百科全书(Kyoto encyclopedia of genes and genomes, KEGG)分析进一步揭示其可能通过调控细胞周期、肾素-血管紧张素系统等多条通路发挥生物学功能。值得注意的是，差异蛋白质中HMOX1、TFRC等低氧应答蛋白质表达上调，而肾素-血管紧张素系统相关通路受到抑制。本研究首次系统阐明了低氧预处理对猫ADMSCs来源外泌体蛋白质组的重塑作用，为深入理解外泌体介导的细胞间通讯提供了新的理论基础。.

Industrial wastewater from battery manufacturing, metal smelting, and electroplating often contains high concentrations of cadmium and nitrogen, resulting in cadmium-nitrogen combined pollution. Conventional physicochemical methods for simultaneous removal of cadmium and nitrogen face limitations such as complex processes, high energy consumption, and risks of secondary pollution. In contrast, using cadmium-tolerant denitrifying microbes, which achieve synergistic removal of cadmium and nitrogen through adsorption, biomineralization, and other pathways, offers advantages of low energy consumption and high efficiency. In this review, we summarize the classification and denitrification characteristics of cadmium-tolerant denitrifying microbes, with a focus on the cadmium tolerance mechanisms and nitrogen removal performance of Pseudomonas as a representative microbial group. We analyze the inhibitory effects of cadmium on denitrification, explore the underlying molecular mechanisms, and summarize microbial response mechanisms and cadmium removal pathways under cadmium stress. In addition, we review the advances in the application of functional bacterial strains in various reactor systems for synergistic cadmium and nitrogen removal, and reviews the achievements in enhancing removal efficiency and system stability through biomineralization and carrier optimization. Finally, we discuss current challenges and limitations and makes an outlook on the future research directions, aiming to provide theoretical and practical insights for the bioremediation of wastewater co-contaminated with cadmium and nitrogen. 在电池生产、金属冶炼和电镀等工业生产过程中产生的废水同时含有高浓度镉和氮，会形成镉-氮复合污染。针对该类废水中同步去除镉、氮的难题，传统物理化学处理方法存在工艺复杂、能耗高、易产生二次污染等局限。相比之下，利用耐镉脱氮微生物，通过吸附、生物矿化等途径实现镉-氮协同去除，表现出低能耗与高效性的优势。本文综述了耐镉脱氮微生物的分类与脱氮特性，重点分析了以假单胞菌属为代表的该类微生物的耐镉机制及脱氮性能;系统分析了镉对脱氮过程的抑制效应及其分子机制;归纳了微生物在镉胁迫下的响应机制和除镉途径;并概述了功能菌株在不同反应工艺中实现镉-氮协同去除的研究进展，总结了生物膜与固定化床反应器通过生物矿化及复合载体构建在提升污染物去除效率与系统稳定性方面取得的成果。最后，基于研究现状分析了存在的问题与局限性，并对未来的研究方向进行了展望，为镉-氮复合污染废水的生物治理提供了理论与实践参考。.

After China implemented the policy of banning the application of antibiotics in feed and reducing the application of antibiotics in animal production, there has been an urgent need to develop new antibiotic alternatives. In this context, animal venom peptides are considered highly promising alternatives to antibiotics because they possess multifunctional properties and appear to employ the same membrane-targeting mechanism for both insecticidal and bactericidal activities. This study examines the antibacterial activity of U10-MYRTX-Mri1a, an insecticidal peptide derived from the venom of European red ants (Manica rubida), and assesses the functionality of its recombinant fusion protein produced in Pichia pastoris, thereby providing experimental support for the development of antibiotic alternatives. According to the concept of structural similarity-functional crossover, we evaluated the antibacterial activities of the insecticidal peptide U10-MYRTX-Mri1a derived from Myrmica rubra against Escherichia coli O157:H7 and Staphylococcus aureus ATCC 25923. U10-MYRTX-Mri1a was chemically synthesized. Its minimum inhibitory concentrations (MIC) were 16 µmol/L for E. coli O157:H7 and 8 µmol/L for S. aureus ATCC 25923, and the minimum bactericidal concentrations for both strains were 16 µmol/L. Scanning electron microscopy showed that the peptide treatment caused collapse of the bacterial membrane structure and cell fragmentation. Meanwhile, the fusion peptide U10-MYRTX-Mri1a-GNA was expressed via the constitutive promoter glyceraldehyde-3-phosphate dehydrogenase (GAP) in Pichia pastoris X33. The fusion peptide inhibited E. coli O157:H7 and S. aureus ATCC 25923 by (89.37±2.22)% and (88.27±2.81)%, respectively. The findings indicate that the insecticidal peptide U10-MYRTX-Mri1a exhibits promising potential as a natural antibiotic substitute, thereby broadening the spectrum of available antibiotic-alternative agents. Furthermore, a reliable biosynthesis platform using Pichia pastoris was successfully developed, which lays a solid groundwork for future research and development of antibiotic alternatives. 我国“饲料禁抗、养殖减抗”政策实施后，新型抗生素替代产品的研发已成为迫切需求。在这种背景下，杀虫肽因具有功能多效性特征，且其杀虫活性与抗菌活性可能共享膜靶向机制，被认为是极具潜力的抗生素替代产品。本研究旨在验证源自欧洲红蚁毒液的杀虫肽U10-MYRTX-Mri1a的抗菌活性，并建立基于毕赤酵母表达系统的该肽生物合成平台，为开发抗生素替代产品提供实验依据与技术支持。基于“结构相似-功能跨界”的思路，本研究验证了U10-MYRTX-Mri1a对大肠杆菌和金黄色葡萄球菌的抗菌活性。通过化学合成获得U10-MYRTX-Mri1a，测定其对2种致病菌的最小抑菌浓度(minimum inhibitory concentration, MIC)分别为16 µmol/L和8 µmol/L，最小杀菌浓度(minimum bactericidal concentration, MBC)均为16 µmol/L。扫描电镜观察到肽处理导致菌体膜结构塌陷和细胞碎裂。同时，本研究采用组成型启动子甘油醛-3-磷酸脱氢酶(glyceraldehyde-3-phosphate dehydrogenase, GAP)，在毕赤酵母X33中表达融合肽U10-MYRTX-Mri1a-GNA，该融合肽对大肠杆菌和金黄色葡萄球菌的抑菌率分别为(89.37±2.22)%、(88.27±2.81)%，证明酵母表达的融合肽同样具有抗菌活性。上述研究结果初步证明杀虫肽U10-MYRTX-Mri1a是有抗生素替代潜力的天然产物，该结果拓宽了替抗产品来源。同时，成功建立的基于毕赤酵母生物合成肽平台，也为后续替抗研究与开发奠定了基础。.

O-acetyl- l- homoserine (OAH) is a key intermediate in the synthesis of various high-value compounds such as l-methionine and has a high market value. However, the production of OAH still faces problems such as low yields and long fermentation periods. In this study, the wild type of the probiotic Escherichia coli Nissle 1917 (EcN) was used as the starting strain. Firstly, we obtained the mutant MetXK185R-G210S-N204T by modifying the key enzyme l-homoserine acetyltransferase (MetX). Further, ppc, asd, and metL were overexpressed to strengthen the synthetic pathways of the precursors l-aspartic acid and l-homoserine. Strategies such as dynamic regulation were adopted to weaken the synthesis of the by-product amino acids. Different intensities of 5{L-End} ' UTR were screened to downregulate the expression level of gltA to balance the supply of another precursor acetyl-CoA, and finally an efficient OAH-producing strain was constructed. The fermentation yield of OAH in the shake flask was 11.18 g/L, and that in the 5 L fermenter reached 63.54 g/L at the time point of 52 h. The strain engineering improved the yield and productivity of OAH and laid a research foundation for the subsequent biological production of l-methionine. O-乙酰- l-高丝氨酸(O-acetyl- l-homoserine, OAH)是 l-蛋氨酸等多种高附加值化合物合成的关键中间体，具有较高的市场价值，但目前OAH生产仍面临产量低、发酵周期长等问题。本研究以大肠杆菌益生菌Escherichia coli Nissle 1917 (EcN)野生型为出发菌株，首先通过对关键合成酶—— l -高丝氨酸乙酰转移酶(l-homoserine O-acetyltransferase, MetX)的改造，获得突变体MetXK185R-G210S-N204T;进一步过表达ppc、asd、metL等基因，强化了前体 l-天冬氨酸和 l-高丝氨酸的合成途径;采用动态调控等策略减弱了副产物氨基酸的合成;并通过筛选不同强度5{L-End} ' UTR，下调gltA基因表达水平，平衡了另一前体乙酰辅酶A的供应，构建了OAH高效生产菌株。最终菌株摇瓶中OAH发酵产量提升至11.18 g/L，5 L发酵罐产量在52 h达到63.54 g/L，本研究提高了OAH的产量和生产效率，为后续 l-蛋氨酸的生物法生产奠定了良好的研究基础。.

To address the delayed heating and insufficient thermophilic phase caused by early mass transfer limitations and local oxygen shortage in aerobic composting, this study investigated the impacts of electric field intervention on the microbial ecological network and functional phenotypes. Composting experiments were conducted in 30 L reactors under different electric field intensities (0 V, 2 V, and 5 V). Physicochemical measurements, 16S rRNA-based community profiling, co-occurrence network analysis, and BugBase phenotype predictions were integrated to elucidate the coupling mechanism of electric field-microorganisms-process. The results demonstrated that electric field application markedly accelerated temperature rise and extended the thermophilic phase, with early-stage current dynamics tightly coupled to temperature. Although temporal succession remained the primary driver of microbial community structure, electric field treatments exerted significant additional effects, resulting in distinct community structures on days 7 and 18. At the genus level, electric field application enriched key taxa associated with electron transfer and fermentation (e.g., Pseudomonas, Proteiniphilum, Bacteroides, Methanocorpusculum, and Peptococcus). The network structure shifted from a "low-aggregation, short-path" pattern to a "high-aggregation, long-path" configuration, with the electric field reshaping the module affiliations and roles of dominant taxa and connectors. Weak fields promoted cross-module cooperation, whereas strong fields enhanced module compartmentalization and hub connectivity. The phenotypic level of BugBase showed periodic migration, and the 5-V electric field reshaped the bacteria to predict the phenotype spectrum effect stronger and earlier. The electric field treatment group was more inclined to enrich the phenotypes of aerobic bacteria, Gram-negative bacteria, biofilm formation, and stress tolerance, while tending to decrease Gram-positive phenotype. Collectively, the electric field enhanced microscale mass transfer and electron acceptor availability, thereby intensifying early metabolic and exothermic processes and restructuring the microbial network, achieving a state of acceleration without directional change. This study elucidates the microbial ecological mechanisms underlying electric-field regulation of aerobic composting, thereby providing a theoretical basis and technical framework for physical-field-enabled, precision modulation of microbial community functions and for the targeted optimization of composting processes. 为缓解好氧堆肥早期传质受限与局部缺氧导致的升温滞后与高温期不足，厘清电场介入对微生物生态网络与功能表型的影响，本研究设置不同强度(0 V、2 V、5 V)的电场施加于30 L反应器的堆肥过程，结合理化指标、16S rRNA细菌群落结构分析、共现网络与BugBase表型预测，解析电场堆肥下微生物群落组成与功能演替机制。研究结果显示:电场能够显著加速升温并延长高温期，前期电流与温度呈紧密耦合;堆肥时间是细菌群落结构的主要影响因素，但在控制时间后，电场效应依旧显著，且在第7天和第18天形成了显著分化的群落结构。属水平上，电场富集了与电子转移/发酵相关的关键类群，例如假单胞菌属(Pseudomonas)、嗜蛋白质菌属(Proteiniphilum)、拟杆菌属(Bacteroides)、甲烷粒菌属(Methanocorpusculum)、消化球菌属(Peptococcus)等，富集了发酵菌和电活性菌;网络结构由“低聚集、短路径”重组为“高聚集、长路径”，电场改变了优势菌与连接者的模块归属和相对位置，弱电场增强跨模块协同，强电场强化模块分区与枢纽连接。BugBase表型层面呈现阶段性迁移，且5 V电场对细菌表型谱的调控效应更强、更早出现。电场处理组更倾向于富集需氧型、革兰氏阴性、生物膜形成与应激耐受等表型，同时革兰氏阳性表型倾向降低。综上，电场通过改善微尺度传质与电子受体可用性，放大了前期代谢与放热过程，微生物网络结构重组，实现了“加速而不改向”。本研究阐明了电场调控好氧堆肥进程的微生物生态学机制，为通过物理场精准调控菌群功能、定向优化堆肥工艺提供了新的理论与技术路径。.

Sulfonamides (SAs) are frequently detected in natural water bodies, sediments, and wastewater treatment plants (WWTPs). Their persistence in the environment induces the generation and dissemination of antibiotic resistance genes (ARGs) as well as drug-resistant bacteria, posing threats to ecosystems and public health. Current processes are insufficient for the removal of SAs. This paper reviews the recent research progress in SAs in terms of the occurrence concentrations, biological toxicity, and induction of resistance gene transfer, as well as our preliminary work on the degradation technology and mechanisms of SAs. Through the review, this paper summarizes the pollution status, hazards, biodegradation processes and key influencing factors of SAs, clarifies the roles of biological community interactions, microbial co-metabolism, eukaryotic enzymatic degradation, and interactions between gut microbiota and the host in the degradation of SAs and horizontal transfer control process of ARGs, and outlines representative degradation pathways, degradation kinetics, and related research methods. Furthermore, this paper makes an outlook on the future research directions from the aspects of innovative combination process, multi-factor synergistic influencing mechanisms, and mutually beneficial cooperation between eukaryotes and their microbial communities, aiming to provide insights for deciphering the degradation mechanisms of SAs and formulating pollution control measures. 磺胺类抗生素(sulfonamides, SAs)在自然水体、沉积物和污水处理厂中检出率高，长期存在会诱导抗性基因和耐药菌的产生和传播，对生态系统和公共健康构成威胁。我国城镇污水处理厂现有工艺难以实现对SAs的有效削减。本文基于近年来国内外学者在SAs检出浓度、生物毒性、诱导抗性基因转移等方面的研究进展，结合本课题组开展的SAs降解技术与机制研究方面的工作，总结了污水中SAs的污染现状和危害、生物降解工艺、关键影响因素，阐明了生物群落协作、微生物共代谢、真核生物酶解、肠道菌群与宿主互作等在SAs降解和抗性基因(antibiotic resistance genes, ARGs)水平转移控制过程中的作用原理，梳理了代表性SAs的降解路径、降解动力学和相关研究方法，并从组合工艺创新、多因素协同影响机制、真核生物及其微生物组互利协作等方面对未来研究方向进行了展望，为SAs降解机理的深入研究和污染治理提供参考。.