Enzyme and Microbial Technology最新文献_第6页

Enhanced synthesis of alanyl-glutamine dipeptide via molecular modification and process optimization of α-amino acid ester acyltransferase EAET α-氨基酸酯酰基转移酶EAET的分子修饰和工艺优化促进丙氨酰-谷氨酰胺二肽的合成。

IF 3.7 3区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Enzyme and Microbial Technology

Pub Date : 2025-09-22 DOI: 10.1016/j.enzmictec.2025.110754

Jinao Wei , Xiaoxi Liu , Yifei Gao, Jing-wen Yang, Hong-bin Zhang, Xue-qin Hu

This study presents a significant advancement in the enzymatic synthesis of alanyl-glutamine (Ala-Gln) through the development of a high-performance α-ester acyltransferase (EAET) using molecular engineering. We successfully cloned EAET from Bacillus pumilus, which exhibits low sequence identity (<30 %) compared to other reported acyltransferases (AETs). Using AlphaFold3, we constructed a highly reliable structural model of EAET, validated by metrics such as pLDDT, ipTM, and PAE. Through semi-rational design, we introduced the F330Y mutation, which enhanced enzymatic activity by 1.5-fold and extended the half-life at 30°C by 1.3-fold. Molecular dynamics simulations revealed increased flexibility around residue 330 and additional hydrogen bonds in the F330Y mutant, both of which contribute to its improved catalytic efficiency and thermostability. Response surface methodology was employed to optimize enzyme production conditions, resulting in a maximum enzyme activity of 218.20 U/mL[91.3 U/(mL·OD600)]. Systematic optimization of catalytic parameters, including pH, temperature, substrate ratio, concentration, enzyme dosage, and reaction time, enabled an 82.59 % conversion yield at 300 mM substrate concentration. These results not only surpass previous benchmarks but also demonstrate the industrial potential of engineered EAET variants for Ala-Gln synthesis. This study highlights the successful integration of computational tools with molecular engineering to enhance enzyme performance, offering a greener and more sustainable approach for Ala-Gln production.

本研究利用分子工程技术开发了高性能α-酯酰基转移酶（EAET），在酶促合成丙氨酰-谷氨酰胺（Ala-Gln）方面取得了重大进展。我们成功地从矮芽孢杆菌中克隆了EAET，该菌株具有低序列同源性(

{"title":"Enhanced synthesis of alanyl-glutamine dipeptide via molecular modification and process optimization of α-amino acid ester acyltransferase EAET","authors":"Jinao Wei , Xiaoxi Liu , Yifei Gao, Jing-wen Yang, Hong-bin Zhang, Xue-qin Hu","doi":"10.1016/j.enzmictec.2025.110754","DOIUrl":"10.1016/j.enzmictec.2025.110754","url":null,"abstract":"<div><div>This study presents a significant advancement in the enzymatic synthesis of alanyl-glutamine (Ala-Gln) through the development of a high-performance α-ester acyltransferase (EAET) using molecular engineering. We successfully cloned EAET from <em>Bacillus pumilus</em>, which exhibits low sequence identity (<30 %) compared to other reported acyltransferases (AETs). Using AlphaFold3, we constructed a highly reliable structural model of EAET, validated by metrics such as pLDDT, ipTM, and PAE. Through semi-rational design, we introduced the F330Y mutation, which enhanced enzymatic activity by 1.5-fold and extended the half-life at 30°C by 1.3-fold. Molecular dynamics simulations revealed increased flexibility around residue 330 and additional hydrogen bonds in the F330Y mutant, both of which contribute to its improved catalytic efficiency and thermostability. Response surface methodology was employed to optimize enzyme production conditions, resulting in a maximum enzyme activity of 218.20 U/mL[91.3 U/(mL·OD600)]. Systematic optimization of catalytic parameters, including pH, temperature, substrate ratio, concentration, enzyme dosage, and reaction time, enabled an 82.59 % conversion yield at 300 mM substrate concentration. These results not only surpass previous benchmarks but also demonstrate the industrial potential of engineered EAET variants for Ala-Gln synthesis. This study highlights the successful integration of computational tools with molecular engineering to enhance enzyme performance, offering a greener and more sustainable approach for Ala-Gln production.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"192 ","pages":"Article 110754"},"PeriodicalIF":3.7,"publicationDate":"2025-09-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145205964","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

A thermostable cellobiose phosphorylase from Thermoclostridium caenicola and its application to in vitro biotransformation 一种耐热性纤维二糖磷酸化酶及其在体外生物转化中的应用

IF 3.7 3区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Enzyme and Microbial Technology

Pub Date : 2025-09-19 DOI: 10.1016/j.enzmictec.2025.110753

Haotian Cheng , Yuan Li , Yi Zhou , Shuke Wu , Yi-Heng P. Job Zhang

Cellobiose phosphorylase (CBP) catalyzes the reversible phosphorolysis of cellobiose to glucose and α-D-glucose-1-phosphate. CBPs play an important role in the in vivo enzymatic utilization of cellulosic materials. Here we discovered a novel CBP from Thermoclostridium caenicola (TcCBP), displaying 50–75 % sequence homology with previously reported CBPs. Recombinant TcCBP was well expressed in E. coli BL21(DE3), with a 1.6-fold increase in soluble expression levels over the widely-used Clostridium thermocellum CBP (CtCBP). This enzyme exhibited broad pH adaptability, maintaining substantial activities across pH 4.0–7.5 in the synthetic direction and pH 5.0–7.5 in the phosphorolytic direction. Compared to CtCBP, TcCBP displayed the superior thermostability and a nearly 100-fold improvement in "Product-to-Enzyme Ratio" (PE value). The biosynthesis of myo-inositol from cellobiose was conducted by using this CBP along with other four thermophilic enzymes (i.e., phosphoglucomutase, inositol 1-phosphate synthase, inositol monophosphatase, and polyphosphate glucokinase) in one pot without step-by-step addition of enzymes. Approximately 96 mM of myo-inositol was produced from 50 mM of cellobiose. These results indicated that this enzyme could be a potential thermophilic enzyme used for the production of value-added biochemicals by in vitro BioTransformation.

纤维素二糖磷酸化酶（CBP）催化纤维素二糖可逆磷酸解生成葡萄糖和α- d -葡萄糖-1-磷酸。CBPs在纤维素材料的体内酶利用中起着重要作用。在这里，我们发现了一个新的CBP从热梭菌caenicola (TcCBP)，显示50 - 75% %序列同源性与先前报道的CBPs。重组TcCBP在大肠杆菌BL21（DE3）中表达良好，可溶性表达量比广泛使用的热胞梭菌CBP （CtCBP）高1.6倍。该酶表现出广泛的pH适应性，在pH 4.0-7.5的合成方向和pH 5.0-7.5的磷酸化方向上保持大量的活性。与CtCBP相比，TcCBP表现出更好的热稳定性，“产物与酶比”（PE值）提高了近100倍。利用该CBP和其他四种嗜热酶（即磷酸葡萄糖化酶、肌醇1-磷酸合成酶、肌醇单磷酸酶和多磷酸葡萄糖激酶）在一个锅中进行了由纤维素糖合成肌醇的生物合成，而无需逐步添加酶。大约96 mM的肌醇由50 mM的纤维二糖产生。这些结果表明，该酶可能是一种潜在的通过体外生物转化生产增值生化产品的嗜热酶。

{"title":"A thermostable cellobiose phosphorylase from Thermoclostridium caenicola and its application to in vitro biotransformation","authors":"Haotian Cheng , Yuan Li , Yi Zhou , Shuke Wu , Yi-Heng P. Job Zhang","doi":"10.1016/j.enzmictec.2025.110753","DOIUrl":"10.1016/j.enzmictec.2025.110753","url":null,"abstract":"<div><div>Cellobiose phosphorylase (CBP) catalyzes the reversible phosphorolysis of cellobiose to glucose and α-D-glucose-1-phosphate. CBPs play an important role in the <em>in vivo</em> enzymatic utilization of cellulosic materials. Here we discovered a novel CBP from <em>Thermoclostridium caenicola</em> (TcCBP), displaying 50–75 % sequence homology with previously reported CBPs. Recombinant TcCBP was well expressed in <em>E. coli</em> BL21(DE3), with a 1.6-fold increase in soluble expression levels over the widely-used <em>Clostridium thermocellum</em> CBP (CtCBP). This enzyme exhibited broad pH adaptability, maintaining substantial activities across pH 4.0–7.5 in the synthetic direction and pH 5.0–7.5 in the phosphorolytic direction. Compared to CtCBP, TcCBP displayed the superior thermostability and a nearly 100-fold improvement in \"Product-to-Enzyme Ratio\" (PE value). The biosynthesis of <em>myo</em>-inositol from cellobiose was conducted by using this CBP along with other four thermophilic enzymes (i.e., phosphoglucomutase, inositol 1-phosphate synthase, inositol monophosphatase, and polyphosphate glucokinase) in one pot without step-by-step addition of enzymes. Approximately 96 mM of <em>myo</em>-inositol was produced from 50 mM of cellobiose. These results indicated that this enzyme could be a potential thermophilic enzyme used for the production of value-added biochemicals by <em>in vitro</em> BioTransformation.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"192 ","pages":"Article 110753"},"PeriodicalIF":3.7,"publicationDate":"2025-09-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145109831","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Strong alone, weak together: biofilm tensile strength in kangaroo rat burrows 单独强，一起弱：袋鼠大鼠洞中的生物膜拉伸强度

IF 3.7 3区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Enzyme and Microbial Technology

Pub Date : 2025-09-17 DOI: 10.1016/j.enzmictec.2025.110752

Duygu Aydin , Idil Deniz Akin , Douglas R. Call , Haluk Beyenal

Desert kangaroo rats (Dipodomys deserti) construct burrows that create protected micro-niches favorable to increased microbial activity and biofilm formation. Biofilms within these burrows bind sand particles together, increase the tensile strength of the burrow sand and burrow stability. Our previous work 1) demonstrated that kangaroo rat burrow sand exhibits higher tensile strength than surrounding surface sand due to the cementation by biofilms, and 2) characterized the microbial biofilm communities within kangaroo rat burrow sand and isolated abundant microorganisms. In this study, isolated species are used to quantify tensile strength of pure and mixed biofilms using the microcantilever technique. Mono-species biofilms of Aspergillus tamarii (59.30 ± 4.36 kPa) exhibited the highest tensile strength, while Neobacillus niacini (9.45 ± 1.98 kPa) showed the lowest. Dual-species biofilms displayed synergistic or antagonistic effects, depending on species combinations. Mixing N. niacini with Peribacillus frigoritolerans increased tensile strength to 55.11 ± 2.51 kPa, whereas combining A. tamarii with N. niacini reduced the tensile strength to 18.98 ± 2.54 kPa. Mixing up to five species reduced tensile strength to a minimum value of 2.16 kPa. We conclude that biofilms formed by microbial isolates from burrow sand individually had higher tensile strength, but when all were mixed, the tensile strength decreased, making them weaker.

沙漠袋鼠鼠（Dipodomys deserti）建造洞穴，创造有利于增加微生物活性和生物膜形成的受保护的微生态位。这些洞穴中的生物膜将沙子颗粒结合在一起，增加了洞穴沙子的抗拉强度和洞穴的稳定性。我们的前期工作表明：1)由于生物膜的胶结作用，袋鼠鼠洞砂具有比周围表面砂更高的抗拉强度；2)表征了袋鼠鼠洞砂内的微生物生物膜群落，并分离出丰富的微生物。在这项研究中，分离物种被用来量化使用微悬臂技术的纯和混合生物膜的抗拉强度。单种生物膜抗拉强度最高的是柽柳曲霉（59.30 ± 4.36 kPa），最低的是烟酸新杆菌（9.45 ± 1.98 kPa）。双物种生物膜表现出协同或拮抗作用，这取决于物种组合。烟芽孢杆菌与冷冻大芽孢杆菌混合后的抗拉强度可达55.11 ± 2.51 kPa，而tamarii与烟芽孢杆菌混合后的抗拉强度可达18.98 ± 2.54 kPa。混合多达五种物质将抗拉强度降低到最小值2.16 kPa。结果表明，单独形成的生物膜具有较高的抗拉强度，但混合形成的生物膜抗拉强度降低，使生物膜变弱。

{"title":"Strong alone, weak together: biofilm tensile strength in kangaroo rat burrows","authors":"Duygu Aydin , Idil Deniz Akin , Douglas R. Call , Haluk Beyenal","doi":"10.1016/j.enzmictec.2025.110752","DOIUrl":"10.1016/j.enzmictec.2025.110752","url":null,"abstract":"<div><div>Desert kangaroo rats (<em>Dipodomys deserti</em>) construct burrows that create protected micro-niches favorable to increased microbial activity and biofilm formation. Biofilms within these burrows bind sand particles together, increase the tensile strength of the burrow sand and burrow stability. Our previous work 1) demonstrated that kangaroo rat burrow sand exhibits higher tensile strength than surrounding surface sand due to the cementation by biofilms, and 2) characterized the microbial biofilm communities within kangaroo rat burrow sand and isolated abundant microorganisms. In this study, isolated species are used to quantify tensile strength of pure and mixed biofilms using the microcantilever technique. Mono-species biofilms of <em>Aspergillus tamarii</em> (59.30 ± 4.36 kPa) exhibited the highest tensile strength, while <em>Neobacillus niacini</em> (9.45 ± 1.98 kPa) showed the lowest. Dual-species biofilms displayed synergistic or antagonistic effects, depending on species combinations. Mixing <em>N. niacini</em> with <em>Peribacillus frigoritolerans</em> increased tensile strength to 55.11 ± 2.51 kPa, whereas combining <em>A. tamarii</em> with <em>N. niacini</em> reduced the tensile strength to 18.98 ± 2.54 kPa. Mixing up to five species reduced tensile strength to a minimum value of 2.16 kPa. We conclude that biofilms formed by microbial isolates from burrow sand individually had higher tensile strength, but when all were mixed, the tensile strength decreased, making them weaker.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"192 ","pages":"Article 110752"},"PeriodicalIF":3.7,"publicationDate":"2025-09-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145106315","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

De Novo enzyme design of a minimal and soluble carbonic anhydrase from Mesorhizobium loti assisted by molecular dynamics simulation 在分子动力学模拟的辅助下，从loti中分离出一种最小的可溶碳酸酐酶

IF 3.7 3区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Enzyme and Microbial Technology

Pub Date : 2025-09-12 DOI: 10.1016/j.enzmictec.2025.110751

Jiun-Jang Juo, I.-Son Ng

Carbonic anhydrases (CAs) are critical biocatalysts in the carbon capture and utilization due to their remarkable efficiency in converting carbon dioxide into bicarbonate. Among all, the CA from Mesorhizobium loti (MlCA) exhibits the highest catalytic activity. However, the poor expression and low solubility in Escherichia coli significantly restricts its application. To overcome the challenges, we employed a protein minimization strategy to improve both enzymatic expression and solubility. Enhanced expressions were observed in genetic constructs with short N-terminal tags which MlCA expression is strongly affected by mRNA secondary structure near the start codon. For solubility issue, a de novo protein design workflow guided by molecular dynamics simulations was developed. The process consists of four stages: (1) size-constrained de novo design considered using AlphaFold and RFdiffusion, (2) sequence recovery replying on solubleMPNN and ESMfold, (3) In silico screening by FoldX, SASA and molecular dynamics evaluation, and (4) experimental validation. This enabled the generation of compact, stable, and catalytically active MlCA variants with 28 % reduction in protein size among 500 candidates. Finally, the most promising design, dM22, of minimal CA showed improved solubility from 16.1 % to 61.2 % in B7G, a GroELS integrated BL21(DE3) strain.

碳酸酐酶（carbon anhydrase, CAs）具有将二氧化碳转化为碳酸氢盐的显著效率，是碳捕获和利用中的关键生物催化剂。其中，loti中根瘤菌（MlCA）的CA表现出最高的催化活性。但其在大肠杆菌中的低表达和低溶解度严重限制了其应用。为了克服这些挑战，我们采用了蛋白质最小化策略来提高酶的表达和溶解性。在具有短n端标签的遗传结构中，MlCA的表达受到起始密码子附近mRNA二级结构的强烈影响。针对溶解度问题，建立了以分子动力学模拟为指导的蛋白质从头设计流程。该过程包括四个阶段：(1)使用AlphaFold和RFdiffusion考虑尺寸约束的从头设计，(2)依靠solubleMPNN和ESMfold进行序列恢复，(3)通过FoldX， SASA和分子动力学评估进行硅筛选，(4)实验验证。这使得在500个候选基因中产生紧凑、稳定和催化活性的MlCA变异，其蛋白质大小减少了28% %。最后，最有希望的设计dM22，最小CA的设计在GroELS整合BL21（DE3）菌株B7G中的溶解度从16.1 %提高到61.2 %。

{"title":"De Novo enzyme design of a minimal and soluble carbonic anhydrase from Mesorhizobium loti assisted by molecular dynamics simulation","authors":"Jiun-Jang Juo, I.-Son Ng","doi":"10.1016/j.enzmictec.2025.110751","DOIUrl":"10.1016/j.enzmictec.2025.110751","url":null,"abstract":"<div><div>Carbonic anhydrases (CAs) are critical biocatalysts in the carbon capture and utilization due to their remarkable efficiency in converting carbon dioxide into bicarbonate. Among all, the CA from <em>Mesorhizobium loti</em> (MlCA) exhibits the highest catalytic activity. However, the poor expression and low solubility in <em>Escherichia coli</em> significantly restricts its application. To overcome the challenges, we employed a protein minimization strategy to improve both enzymatic expression and solubility. Enhanced expressions were observed in genetic constructs with short N-terminal tags which MlCA expression is strongly affected by mRNA secondary structure near the start codon. For solubility issue, a <em>de novo</em> protein design workflow guided by molecular dynamics simulations was developed. The process consists of four stages: (1) size-constrained <em>de novo</em> design considered using AlphaFold and RFdiffusion, (2) sequence recovery replying on solubleMPNN and ESMfold, (3) <em>In silico</em> screening by FoldX, SASA and molecular dynamics evaluation, and (4) experimental validation. This enabled the generation of compact, stable, and catalytically active MlCA variants with 28 % reduction in protein size among 500 candidates. Finally, the most promising design, dM22, of minimal CA showed improved solubility from 16.1 % to 61.2 % in B7G, a GroELS integrated BL21(DE3) strain.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"192 ","pages":"Article 110751"},"PeriodicalIF":3.7,"publicationDate":"2025-09-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145057156","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Corrigendum to “Influence of a rumen ciliate-derived xylanase on the gut microbiota composition: A potential enzyme for prebiotic applications” [Enzyme Microb. Technol. 190 (2025) 110683] “瘤胃纤毛虫衍生的木聚糖酶对肠道微生物群组成的影响：一种潜在的益生元应用酶”的更正[微生物酶]。技术通报，190 (2025)110683 [j]。

IF 3.7 3区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Enzyme and Microbial Technology

Pub Date : 2025-09-10 DOI: 10.1016/j.enzmictec.2025.110749

Weide Su , Huiying Luo , Gaoxiang Ai , Qipeng Wei , Zhiheng Zou , Xiaolian Chen , Chuanhui Xu , Jiang Chen , Pingwen Xiong , Wenjing Song , Qiongli Song

引用次数: 0

Homologous expression and characterization of Coniochaeta ligniaria glycoside hydrolase family 115 α-glucuronidase 木质素螺毛菌糖苷水解酶家族115 α-葡萄糖醛酸酶的同源表达与鉴定

IF 3.7 3区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Enzyme and Microbial Technology

Pub Date : 2025-09-09 DOI: 10.1016/j.enzmictec.2025.110750

Michael J. Bowman, Nancy N. Nichols

Generation of fermentable sugars from biomass is necessary for microbial production of bioproducts. Due to its inherent complexity, xylan hydrolysis to monosaccharides requires several different enzymatic activities. To completely depolymerize glucuronoarabinoxylan from biomass, α-glucuronidase activity is necessary. The ascomycete fungus Coniochaeta ligniaria has the ability to degrade xylan and is an unutilized source of biomass degrading enzymes. Therefore, a gene encoding a putative Coniochaeta ligniaria GH115 α-glucuronidase was cloned and expressed homologously in Coniochaeta ligniaria as a native secreted protein. Culture supernatants were concentrated and purified by a combination of ultrafiltration, anion-exchange, and size-exclusion chromatography. The purified protein behaved as dominantly dimeric complexes as determined by size-exclusion chromatography. The expressed protein liberated: 4-O-methyl glucuronic acid from beech xylan, birch xylan, and beech-derived glucuronoxylooligosaccharides as the sole product of hydrolysis; and 4-O-methyl glucuronic acid and glucuronic acid from oat spelt xylan. The expressed α-1,2-glucuronidase had greater activity on glucuronoxylooligosaccharides than on full length beech xylan. The expressed α-1,2-glucuronidase, herein designated ClAgu115, had K_m values of: 1.3 mM; 1.2 mM; and 1.0 mM for beech xylan, GH10-hydrolyzed glucuronooligosaccharides, and GH11-hydrolyzed glucuronooligosaccharides, respectively. The measured kinetic constants show that the enzyme prefers an oligosaccharide substrate with the 4-O-methyl glucuronic acid on the non-reducing end. The enzyme had activity between pH 3.0–6.0 and temperatures 10°C-60°C, with optima at pH 4.3 and 40°C. The expression and characterization of ClAgu115 expands the repertoire of fungal GH115 enzymes for use in biomass conversion.

从生物质中产生可发酵糖是微生物生产生物产品所必需的。由于其固有的复杂性，木聚糖水解成单糖需要几种不同的酶活性。为了从生物质中完全解聚葡萄糖醛酸阿拉伯木聚糖，α-葡萄糖醛酸酶活性是必需的。子囊菌木质素Coniochaeta ligniaria具有降解木聚糖的能力，是未利用的生物质降解酶的来源。因此，我们克隆了一个推测为木质素毛刀菌GH115 α-葡糖醛酸酶的基因，并将其作为天然分泌蛋白在木质素毛刀菌中同源表达。培养上清通过超滤、阴离子交换和尺寸排除层析的组合进行浓缩和纯化。纯化后的蛋白主要表现为二聚体复合物，通过尺寸排除层析确定。所表达的蛋白作为水解的唯一产物，从山毛榉木聚糖、桦木聚糖和山毛榉衍生的低聚葡萄糖醛酸中释放出4- o -甲基葡萄糖醛酸；4- o -甲基葡萄糖醛酸和4- o -甲基葡萄糖醛酸来自燕麦斯佩尔木聚糖。α-1,2-葡萄糖醛酸酶对低聚葡萄糖醛酸的活性高于对全长山毛榉木聚糖的活性。表达的α-1,2-葡糖醛酸酶，本文命名为ClAgu115，其Km值为：1.3 mM； 1.2毫米;山毛榉木聚糖、gh10水解的低聚葡萄糖醛酸糖和gh11水解的低聚葡萄糖醛酸糖分别为1.0 mM。测定的动力学常数表明，酶倾向于低聚糖底物，在非还原端有4- o -甲基葡萄糖醛酸。该酶在pH 3.0 ~ 6.0、温度10℃~ 60℃范围内具有活性，pH 4.3和温度40℃时活性最高。ClAgu115的表达和鉴定扩大了真菌GH115酶用于生物质转化的范围。

{"title":"Homologous expression and characterization of Coniochaeta ligniaria glycoside hydrolase family 115 α-glucuronidase","authors":"Michael J. Bowman, Nancy N. Nichols","doi":"10.1016/j.enzmictec.2025.110750","DOIUrl":"10.1016/j.enzmictec.2025.110750","url":null,"abstract":"<div><div>Generation of fermentable sugars from biomass is necessary for microbial production of bioproducts. Due to its inherent complexity, xylan hydrolysis to monosaccharides requires several different enzymatic activities. To completely depolymerize glucuronoarabinoxylan from biomass, α-glucuronidase activity is necessary. The ascomycete fungus <em>Coniochaeta ligniaria</em> has the ability to degrade xylan and is an unutilized source of biomass degrading enzymes. Therefore, a gene encoding a putative <em>Coniochaeta ligniaria</em> GH115 α-glucuronidase was cloned and expressed homologously in <em>Coniochaeta ligniaria</em> as a native secreted protein<em>.</em> Culture supernatants were concentrated and purified by a combination of ultrafiltration, anion-exchange, and size-exclusion chromatography. The purified protein behaved as dominantly dimeric complexes as determined by size-exclusion chromatography. The expressed protein liberated: 4-<em>O</em>-methyl glucuronic acid from beech xylan, birch xylan, and beech-derived glucuronoxylooligosaccharides as the sole product of hydrolysis; and 4-<em>O</em>-methyl glucuronic acid and glucuronic acid from oat spelt xylan. The expressed α-1,2-glucuronidase had greater activity on glucuronoxylooligosaccharides than on full length beech xylan. The expressed α-1,2-glucuronidase, herein designated <em>Cl</em>Agu115, had <em>K</em><sub><em>m</em></sub> values of: 1.3 mM; 1.2 mM; and 1.0 mM for beech xylan, GH10-hydrolyzed glucuronooligosaccharides, and GH11-hydrolyzed glucuronooligosaccharides, respectively. The measured kinetic constants show that the enzyme prefers an oligosaccharide substrate with the 4-<em>O</em>-methyl glucuronic acid on the non-reducing end. The enzyme had activity between pH 3.0–6.0 and temperatures 10°C-60°C, with optima at pH 4.3 and 40°C. The expression and characterization of <em>Cl</em>Agu115 expands the repertoire of fungal GH115 enzymes for use in biomass conversion.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"192 ","pages":"Article 110750"},"PeriodicalIF":3.7,"publicationDate":"2025-09-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145046785","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Rosmarinic acid production using advanced metabolic engineering strategies 利用先进的代谢工程策略生产迷迭香酸

IF 3.7 3区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Enzyme and Microbial Technology

Pub Date : 2025-09-08 DOI: 10.1016/j.enzmictec.2025.110748

Beining Wang , Jintao Lu , Ruzhe Zhang , Jung-Kul Lee , Vipin Chandra Kalia , Chunjie Gong

With improved socio-economic conditions and heightened health awareness, modern consumers now prioritize the nutritional and functional attributes of food over basic satiety. Recently, a high value component, rosmarinic acid synthesized in plants have gained attention as functional food ingredients. Traditional strategy of rosmarinic acid production, including chemical synthesis and plant extraction, are limited by environmental concerns, low yields, and high costs. With the development of biotechnology, metabolic engineering is an alternative strategy for the efficient and cost-effective production of rosmarinic acid. This review focuses on metabolic engineering advances featuring three core strategies: dynamic pathway regulation, cofactor recycling, and microbial co-culture systems. These potential innovations hold great promise for significantly enhancing rosmarinic acid yields. In addition, the review evaluates the economic and technical feasibility of large-scale production, emphasizing the addressing of challenges from traditional production methods.

随着社会经济条件的改善和健康意识的提高，现代消费者现在优先考虑食品的营养和功能属性，而不是基本的饱腹感。近年来，从植物中合成的高价值成分迷迭香酸作为功能性食品原料受到了广泛关注。传统的迷迭香酸生产策略，包括化学合成和植物提取，受到环境问题、低产量和高成本的限制。随着生物技术的发展，代谢工程是高效、低成本生产迷迭香酸的替代策略。本文综述了代谢工程的三个核心策略：动态途径调控、辅因子循环和微生物共培养系统。这些潜在的创新技术有望显著提高迷迭香酸的产量。此外，本文还评估了大规模生产的经济和技术可行性，强调解决传统生产方法带来的挑战。

{"title":"Rosmarinic acid production using advanced metabolic engineering strategies","authors":"Beining Wang , Jintao Lu , Ruzhe Zhang , Jung-Kul Lee , Vipin Chandra Kalia , Chunjie Gong","doi":"10.1016/j.enzmictec.2025.110748","DOIUrl":"10.1016/j.enzmictec.2025.110748","url":null,"abstract":"<div><div>With improved socio-economic conditions and heightened health awareness, modern consumers now prioritize the nutritional and functional attributes of food over basic satiety. Recently, a high value component, rosmarinic acid synthesized in plants have gained attention as functional food ingredients. Traditional strategy of rosmarinic acid production, including chemical synthesis and plant extraction, are limited by environmental concerns, low yields, and high costs. With the development of biotechnology, metabolic engineering is an alternative strategy for the efficient and cost-effective production of rosmarinic acid. This review focuses on metabolic engineering advances featuring three core strategies: dynamic pathway regulation, cofactor recycling, and microbial co-culture systems. These potential innovations hold great promise for significantly enhancing rosmarinic acid yields. In addition, the review evaluates the economic and technical feasibility of large-scale production, emphasizing the addressing of challenges from traditional production methods.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"192 ","pages":"Article 110748"},"PeriodicalIF":3.7,"publicationDate":"2025-09-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145027364","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Innovative nanosensing methods for lysozyme identification 溶菌酶鉴定的创新纳米传感方法

IF 3.7 3区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Enzyme and Microbial Technology

Pub Date : 2025-09-02 DOI: 10.1016/j.enzmictec.2025.110745

Mohammad Darvishi , Seyyed Mohammad Mousavinia , Reza Soleimany , Mohammad Mahdi Heidari , Mohsen Mohammadi , Samad Rastmanesh , Ahmad Mobed

Lysozyme (LYZ) is a critical enzyme recognized for its significant antimicrobial properties, playing an integral role in the immune response and being implicated in various diseases, including infections and inflammatory conditions. Traditional detection methods for LYZ, such as enzyme-linked immunosorbent assays (ELISA) and spectrophotometric techniques, often encounter limitations regarding sensitivity, specificity, and time efficiency. In light of these challenges, there has been substantial advancement in the development of novel biosensor technologies over the past two decades, particularly those that incorporate nanomaterials. These innovative biosensors demonstrate enhanced performance, facilitating rapid and accurate detection of LYZ at low concentrations. This paper aims to provide a comprehensive overview of recently developed biosensors specifically for LYZ, highlighting their design, functionality, and applications in clinical diagnostics and research. Unlike previous reviews, we place a distinct emphasis on the clinical importance of LYZ and its role in various diseases, thereby elucidating its significance in health and disease contexts. Additionally, we will explore the implications of advanced detection methods, particularly those utilizing nanomaterials, for enhancing our understanding of LYZ-related pathologies. By focusing on nanoscale detection techniques, which have not been thoroughly addressed in existing literature, we underscore the transformative potential of these biosensors in LYZ detection. This approach not only contributes to improved disease management but also informs therapeutic strategies, setting our review apart from prior works.

溶菌酶（LYZ）是一种重要的酶，因其显著的抗菌特性而被公认，在免疫反应中起着不可或缺的作用，并与包括感染和炎症在内的各种疾病有关。传统的LYZ检测方法，如酶联免疫吸附试验（ELISA）和分光光度法技术，经常遇到灵敏度、特异性和时间效率方面的限制。鉴于这些挑战，在过去的二十年里，新型生物传感器技术的发展取得了实质性的进步，特别是那些结合纳米材料的技术。这些创新的生物传感器表现出更高的性能，有助于在低浓度下快速准确地检测LYZ。本文旨在全面概述最近开发的LYZ生物传感器，重点介绍它们的设计，功能和在临床诊断和研究中的应用。与以前的综述不同，我们特别强调LYZ的临床重要性及其在各种疾病中的作用，从而阐明其在健康和疾病环境中的重要性。此外，我们将探讨先进的检测方法的意义，特别是那些利用纳米材料，以提高我们对lyz相关病理的理解。通过关注纳米级检测技术，这在现有文献中尚未得到彻底解决，我们强调了这些生物传感器在LYZ检测中的变革潜力。这种方法不仅有助于改善疾病管理，而且为治疗策略提供信息，使我们的综述与先前的工作不同。

{"title":"Innovative nanosensing methods for lysozyme identification","authors":"Mohammad Darvishi , Seyyed Mohammad Mousavinia , Reza Soleimany , Mohammad Mahdi Heidari , Mohsen Mohammadi , Samad Rastmanesh , Ahmad Mobed","doi":"10.1016/j.enzmictec.2025.110745","DOIUrl":"10.1016/j.enzmictec.2025.110745","url":null,"abstract":"<div><div>Lysozyme (LYZ) is a critical enzyme recognized for its significant antimicrobial properties, playing an integral role in the immune response and being implicated in various diseases, including infections and inflammatory conditions. Traditional detection methods for LYZ, such as enzyme-linked immunosorbent assays (ELISA) and spectrophotometric techniques, often encounter limitations regarding sensitivity, specificity, and time efficiency. In light of these challenges, there has been substantial advancement in the development of novel biosensor technologies over the past two decades, particularly those that incorporate nanomaterials. These innovative biosensors demonstrate enhanced performance, facilitating rapid and accurate detection of LYZ at low concentrations. This paper aims to provide a comprehensive overview of recently developed biosensors specifically for LYZ, highlighting their design, functionality, and applications in clinical diagnostics and research. Unlike previous reviews, we place a distinct emphasis on the clinical importance of LYZ and its role in various diseases, thereby elucidating its significance in health and disease contexts. Additionally, we will explore the implications of advanced detection methods, particularly those utilizing nanomaterials, for enhancing our understanding of LYZ-related pathologies. By focusing on nanoscale detection techniques, which have not been thoroughly addressed in existing literature, we underscore the transformative potential of these biosensors in LYZ detection. This approach not only contributes to improved disease management but also informs therapeutic strategies, setting our review apart from prior works.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"192 ","pages":"Article 110745"},"PeriodicalIF":3.7,"publicationDate":"2025-09-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144989476","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Isolation, cloning, and characterization of a novel GH5 cellulase from yak rumen metagenome for enhanced lignocellulose hydrolysis in biofuel production and ruminant feed utilization 牦牛瘤胃宏基因组中新型GH5纤维素酶的分离、克隆和特性分析，用于提高生物燃料生产和反刍动物饲料利用中的木质纤维素水解

IF 3.7 3区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Enzyme and Microbial Technology

Pub Date : 2025-08-19 DOI: 10.1016/j.enzmictec.2025.110737

Ibrahim Bature , Zeyi Liang , Wu Xiaohu , Feng Yang , Yayuan Yang , Pengcheng Dong , Xuezhi Ding

Lignocellulosic biomass is a promising feedstock for biofuel production, but its complex structure, including cellulose and hemicellulose, challenges efficient enzymatic hydrolysis. CelyA, a novel cellulase from the yak rumen, has potential for improving biomass degradation and biofuel production. The CelyA gene was cloned, expressed, and purified. Biochemical characterization included assessments of pH, temperature, and salt tolerance. The enzyme's substrate specificity was tested on crystalline cellulose, CMC, and other polysaccharides. In vitro rumen fermentation was performed to evaluate its effect on fiber digestibility and microbial community composition. Biofuel production was tested by hydrolyzing maize, rice, and wheat straw. CelyA demonstrated optimal activity at pH 6.0 and 40.0 °C and maintained moderate stability across a wide pH range (3.0–12.0), retaining measurable activity even under strongly acidic and alkaline conditions. The enzyme demonstrated excellent salt tolerance, retaining 91.0 % activity in 1.0 M NaCl. CelyA efficiently degraded maize straw in hydrolysis assays, producing 7.2 µmol/L of reducing sugars. In vitro rumen fermentation with CelyA increased fiber digestibility by 8.3 % for maize straw, 14.5 % for rice straw, and 2.7 % for wheat straw. Gas production also increased significantly, with maize straw showing a 91.3 % increase. 16S rRNA sequencing revealed selective enrichment of Ruminococcus and Prevotella, key cellulolytic microbes. CelyA demonstrates strong potential for biofuel production, efficiently hydrolyzing lignocellulosic biomass and enhancing ruminal fiber digestibility. Its stability, salt tolerance, and substrate specificity make it a valuable enzyme for biofuel production and livestock feed optimization.

木质纤维素生物质是一种很有前途的生物燃料原料，但其复杂的结构，包括纤维素和半纤维素，挑战了高效的酶解。CelyA是一种来自牦牛瘤胃的新型纤维素酶，具有改善生物质降解和生物燃料生产的潜力。对CelyA基因进行克隆、表达和纯化。生化表征包括pH值、温度和耐盐性的评估。该酶对结晶纤维素、CMC和其他多糖的底物特异性进行了测试。通过体外瘤胃发酵研究其对纤维消化率和微生物群落组成的影响。生物燃料的生产是通过水解玉米、水稻和麦秆来测试的。CelyA在pH 6.0和40.0°C时表现出最佳活性，并在较宽的pH范围（3.0-12.0）内保持适度的稳定性，即使在强酸性和碱性条件下也保持可测量的活性。该酶表现出优异的耐盐性，在1.0 M NaCl中保持91.0 %的活性。在水解实验中，CelyA能有效地降解玉米秸秆，产生7.2µmol/L的还原糖。在体外瘤胃发酵条件下，玉米秸秆、水稻秸秆和小麦秸秆的纤维消化率分别提高了8.3 %、14.5 %和2.7 %。产气量也显著增加，其中玉米秸秆增产91.3 %。16S rRNA测序结果显示，Ruminococcus和Prevotella选择性富集，这是关键的纤维素分解微生物。CelyA在生物燃料生产方面具有强大的潜力，可以有效地水解木质纤维素生物质并提高瘤胃纤维的消化率。它的稳定性、耐盐性和底物特异性使其成为生物燃料生产和牲畜饲料优化的重要酶。

{"title":"Isolation, cloning, and characterization of a novel GH5 cellulase from yak rumen metagenome for enhanced lignocellulose hydrolysis in biofuel production and ruminant feed utilization","authors":"Ibrahim Bature , Zeyi Liang , Wu Xiaohu , Feng Yang , Yayuan Yang , Pengcheng Dong , Xuezhi Ding","doi":"10.1016/j.enzmictec.2025.110737","DOIUrl":"10.1016/j.enzmictec.2025.110737","url":null,"abstract":"<div><div>Lignocellulosic biomass is a promising feedstock for biofuel production, but its complex structure, including cellulose and hemicellulose, challenges efficient enzymatic hydrolysis. <em>CelyA</em>, a novel cellulase from the yak rumen, has potential for improving biomass degradation and biofuel production. The <em>CelyA</em> gene was cloned, expressed, and purified. Biochemical characterization included assessments of pH, temperature, and salt tolerance. The enzyme's substrate specificity was tested on crystalline cellulose, CMC, and other polysaccharides. <em>In vitro</em> rumen fermentation was performed to evaluate its effect on fiber digestibility and microbial community composition. Biofuel production was tested by hydrolyzing maize, rice, and wheat straw. <em>CelyA</em> demonstrated optimal activity at pH 6.0 and 40.0 °C and maintained moderate stability across a wide pH range (3.0–12.0), retaining measurable activity even under strongly acidic and alkaline conditions. The enzyme demonstrated excellent salt tolerance, retaining 91.0 % activity in 1.0 M NaCl. <em>CelyA</em> efficiently degraded maize straw in hydrolysis assays, producing 7.2 µmol/L of reducing sugars. <em>In vitro</em> rumen fermentation with <em>CelyA</em> increased fiber digestibility by 8.3 % for maize straw, 14.5 % for rice straw, and 2.7 % for wheat straw. Gas production also increased significantly, with maize straw showing a 91.3 % increase. 16S rRNA sequencing revealed selective enrichment of <em>Ruminococcus</em> and <em>Prevotella</em>, key cellulolytic microbes. <em>CelyA</em> demonstrates strong potential for biofuel production, efficiently hydrolyzing lignocellulosic biomass and enhancing ruminal fiber digestibility. Its stability, salt tolerance, and substrate specificity make it a valuable enzyme for biofuel production and livestock feed optimization.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"191 ","pages":"Article 110737"},"PeriodicalIF":3.7,"publicationDate":"2025-08-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144907276","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Developing a urea biosensor and safe blood cleaning method utilizing a novel high throughput La-doped CeO2 nanosized artificial urease with high biocompatibility and enzyme-like activity 利用具有高生物相容性和酶样活性的新型高通量la掺杂CeO2纳米人工脲酶开发尿素生物传感器和安全血液清洁方法

IF 3.7 3区生物学 Q2 BIOTECHNOLOGY & APPLIED MICROBIOLOGY

Enzyme and Microbial Technology

Pub Date : 2025-08-15 DOI: 10.1016/j.enzmictec.2025.110736

Naushad Ahmad , Ashok Kumar Bishoyi , Suhas Ballal , Aman Shankhyan , Shaker Al-Hasnaawei , Karthikeyan Jayabalan , Laxmidhar Maharana , Madan Lal

Herein, a urea biosensor and safe blood cleaning method utilizing a novel high throughput La-doped CeO₂ nanosized artificial urease with high biocompatibility and enzyme-like activity were developed. The urease-like activity, stability composition, morphological characteristics, size, biocompatibility, and crystalline characteristics of the artificial urease were assessed. Considering its high urease-like activity, it is applied for both urea biosensing and safe blood cleaning. The effective factors on the sensing conditions were optimized, providing a wide linear range over 1–10 µM and a low detection limit of 0.5 µM. The repeatability and reproducibility assessments revealed a %RSD of 1.7 % and 2.7 %, in order. The selectivity studies and real blood analysis revealed highly selective, accurate, and reliable determination of urea in biosamples using the developed method, providing a %recovery of 97.8–99.5 % with a low %RSD range of 1.4–3.3 %. Besides, the safe blood cleaning experiments were also performed by urea mineralization from blood. The results exhibited a high blood cleaning yield of 99.5 % at a short time of 30 min, revealing 1.8-fold higher cleaning efficiency for the artificial urease compared to native urease. The reusability studies showed that the blood cleaning yield was saved for about 10 cycles and then it decreased by increasing the operational cycles, reaching 78.6 % after 17 cycles. The shelf-life of the urease was also evaluated within 30 days, revealing a high shelf-stability for the as-prepared nanosized artificial urease. Consequentially, the nanosized artificial urease can be practically applied for both urea detection and blood cleaning in the real world.

本研究利用高生物相容性和酶样活性的新型高通量la掺杂CeO2纳米人工脲酶，开发了一种尿素生物传感器和安全血液清洁方法。对人工脲酶的类脲酶活性、稳定性组成、形态特征、大小、生物相容性和结晶特性进行了评价。由于其具有高的类脲酶活性，因此可用于尿素生物传感和安全血液清洁。对影响传感条件的因素进行了优化，在1-10 µM范围内具有较宽的线性范围，检测限低至0.5 µM。重复性和再现性评估显示，RSD分别为1.7 %和2.7 %。选择性研究和实际血液分析表明，该方法具有较高的选择性、准确性和可靠性，回收率为97.8 ~ 99.5 %，RSD范围为1.4 ~ 3.3 %。此外，还进行了血液尿素矿化的安全血液清洗实验。结果表明，在30 min的短时间内，人工脲酶的血液清洁率高达99.5 %，比天然脲酶的清洁效率高1.8倍。可重复使用性研究表明，血液清洁率可保存约10个循环，然后随着操作周期的增加而降低，17个循环后达到78.6% %。在30天内对脲酶的保质期进行了评估，表明制备的纳米级人工脲酶具有很高的货架稳定性。因此，纳米级人工脲酶可以在现实世界中用于尿素检测和血液清洁。

{"title":"Developing a urea biosensor and safe blood cleaning method utilizing a novel high throughput La-doped CeO2 nanosized artificial urease with high biocompatibility and enzyme-like activity","authors":"Naushad Ahmad , Ashok Kumar Bishoyi , Suhas Ballal , Aman Shankhyan , Shaker Al-Hasnaawei , Karthikeyan Jayabalan , Laxmidhar Maharana , Madan Lal","doi":"10.1016/j.enzmictec.2025.110736","DOIUrl":"10.1016/j.enzmictec.2025.110736","url":null,"abstract":"<div><div>Herein, a urea biosensor and safe blood cleaning method utilizing a novel high throughput La-doped CeO<sub>2</sub> nanosized artificial urease with high biocompatibility and enzyme-like activity were developed. The urease-like activity, stability composition, morphological characteristics, size, biocompatibility, and crystalline characteristics of the artificial urease were assessed. Considering its high urease-like activity, it is applied for both urea biosensing and safe blood cleaning. The effective factors on the sensing conditions were optimized, providing a wide linear range over 1–10 µM and a low detection limit of 0.5 µM. The repeatability and reproducibility assessments revealed a %RSD of 1.7 % and 2.7 %, in order. The selectivity studies and real blood analysis revealed highly selective, accurate, and reliable determination of urea in biosamples using the developed method, providing a %recovery of 97.8–99.5 % with a low %RSD range of 1.4–3.3 %. Besides, the safe blood cleaning experiments were also performed by urea mineralization from blood. The results exhibited a high blood cleaning yield of 99.5 % at a short time of 30 min, revealing 1.8-fold higher cleaning efficiency for the artificial urease compared to native urease. The reusability studies showed that the blood cleaning yield was saved for about 10 cycles and then it decreased by increasing the operational cycles, reaching 78.6 % after 17 cycles. The shelf-life of the urease was also evaluated within 30 days, revealing a high shelf-stability for the as-prepared nanosized artificial urease. Consequentially, the nanosized artificial urease can be practically applied for both urea detection and blood cleaning in the real world.</div></div>","PeriodicalId":11770,"journal":{"name":"Enzyme and Microbial Technology","volume":"191 ","pages":"Article 110736"},"PeriodicalIF":3.7,"publicationDate":"2025-08-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144864722","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0