Biotechnology Letters最新文献

Combining biotechnology with sustainability: feasibility of bioenergy sorghum in generating high-value bioproducts. 结合生物技术与可持续性：生物能源高粱生产高价值生物产品的可行性。

IF 2.1 4区生物学

Biotechnology Letters Pub Date : 2025-08-01 DOI: 10.1007/s10529-025-03623-2

Trine B Andersen, Elliot Braun, Brianna N I Brown, Bjoern Hamberger, Leah Knoor, İlayda Korkmaz, Lucas Reist, Luke Sharpe, Brian Adam McKinley, James O Suggitt, Mitchell A Ticoras, Angel Indibi

引用次数: 0

Gene disruption via a transient hypercompact CRISPR-AsCas12f1 system in Kluyveromyces marxianus. 通过瞬时超紧凑CRISPR-AsCas12f1系统在马氏克卢维酵母中的基因破坏。

IF 2.1 4区生物学

Biotechnology Letters Pub Date : 2025-07-31 DOI: 10.1007/s10529-025-03626-z

Kehui Zhang, Dongmei Wang, Shenglin Hu, Xingjiang Li, Jiong Hong

{"title":"Gene disruption via a transient hypercompact CRISPR-AsCas12f1 system in Kluyveromyces marxianus.","authors":"Kehui Zhang, Dongmei Wang, Shenglin Hu, Xingjiang Li, Jiong Hong","doi":"10.1007/s10529-025-03626-z","DOIUrl":"https://doi.org/10.1007/s10529-025-03626-z","url":null,"abstract":"Kluyveromyces marxianus is an attractive chassis for microbial cell factories due to its rapid growth, thermotolerance, and wide substrate spectrum. However, gene disruption in this organism is challenging primarily due the prevalence of dominant nonhomologous recombination. AsCas12f1, a hypercompact CRISPR-associated protein consisting of 422 amino acids-approximately one-third the size of Cas9 or Cas12a-enables more efficient packaging into delivery vehicles than its larger counterparts. In this study, a gene disruption method using AsCas12f1 was established in K. marxianus through a transient targeting strategy. The integration of tRNA-gRNA into the gRNA construct increased gene disruption efficiency. Additionally, disrupting KmKU70 or KmLIG4 further increased this efficiency, achieving nearly 100%. By combining the disruption of KmKU70 with the AsCas12f1 system, the length of the homologous arm was shortened to 200 bp while maintaining a disruption efficiency of 87.5%. The implementation of the gRNA-tRNA-array system resulted in the successful generation of three single-gene knockout strains from a single transformation, resulting an overall efficiency of 86.4%. This approach leverages the transient transformation of fragments, eliminates the need for extensive time investment in constructing gRNA expression vectors and negates the requirement for the removal of the CRISPR-AsCas12f1 system after gene disruption. This study presents a novel strategy for gene disruption in K. marxianus and demonstrates the applicability of Cas12f in yeast systems.","PeriodicalId":8929,"journal":{"name":"Biotechnology Letters","volume":"47 4","pages":"84"},"PeriodicalIF":2.1,"publicationDate":"2025-07-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144752240","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Paenibacillus barengoltzii: isolation, growth, and characterization of a high dextrinizer α-amylase. 巴氏芽孢杆菌：高糊化剂α-淀粉酶的分离、生长和特性。

IF 2.1 4区生物学

Biotechnology Letters Pub Date : 2025-07-30 DOI: 10.1007/s10529-025-03624-1

Alonso R Poma Ticona, Mário da Silva Neto, Janice Lisboa de Marco, Roberto Castellanos Cabrera, Pedro R Vieira Hamann, Igor Polikarpov, Eliane Ferreira Noronha

{"title":"Paenibacillus barengoltzii: isolation, growth, and characterization of a high dextrinizer α-amylase.","authors":"Alonso R Poma Ticona, Mário da Silva Neto, Janice Lisboa de Marco, Roberto Castellanos Cabrera, Pedro R Vieira Hamann, Igor Polikarpov, Eliane Ferreira Noronha","doi":"10.1007/s10529-025-03624-1","DOIUrl":"https://doi.org/10.1007/s10529-025-03624-1","url":null,"abstract":"Currently, the shift to a greener economy requires the prospection of new industrial processes to reduce greenhouse-gas emissions. Enzymatic catalysis is considered a green alternative to traditional industrial processes. Among enzymes of industrial relevance, starch-degrading enzymes, such as α-amylases, have received attention because of their enormous potential to hydrolyze starch-based materials, generating smaller sugars that can be used for the biosynthesis of chemicals of industrial relevance, such as ethanol. In the present study, a new isolate of Paenibacillus barengoltzii was obtained from cow rumen and its potential to produce amylases was evaluated. Additionally, a recombinant amylase, AmyPb, was produced and biochemically characterized. AmyPb displays high activity at elevated temperatures (55 °C) and can withstand elevated temperatures. The experimentally calculated melting temperature showed that AmyPb is more stable in alkaline environments, with a Tm of 59 °C at pH 9. AmyPb hydrolyzed potato and cassava starches with hydrolysis efficiencies of 28 and 55%, respectively. The results of this study are relevant for the development of industrial processes that employ thermostable amylases.","PeriodicalId":8929,"journal":{"name":"Biotechnology Letters","volume":"47 4","pages":"83"},"PeriodicalIF":2.1,"publicationDate":"2025-07-30","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144752241","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Recombinant protein CP01850 adjuvanted with Iridea cordata or Sacorpletis skottibergii lipid extracts protected mice against infection by Corynebacterium pseudotuberculosis. 重组蛋白CP01850佐剂鸢尾或棘猴脂质提取物对小鼠假结核棒状杆菌感染有保护作用。

IF 2 4区生物学

Biotechnology Letters Pub Date : 2025-07-24 DOI: 10.1007/s10529-025-03622-3

Tallyson Nogueira Barbosa, Nicole Ramos Scholl, Mara Thais de Oliveira Silva, Adriane Leites Strothmann, Henrique Gonçalves Pegoraro, Fernanda Severo Sabedra Sousa, Fabiana Kommling Seixas, Francisco Silvestre Brilhante Bezerra, Tiago Veiras Collares, Cláudio Martin Pereira de Pereira, Andrés Mansilla, Sibele Borsuk

{"title":"Recombinant protein CP01850 adjuvanted with Iridea cordata or Sacorpletis skottibergii lipid extracts protected mice against infection by Corynebacterium pseudotuberculosis.","authors":"Tallyson Nogueira Barbosa, Nicole Ramos Scholl, Mara Thais de Oliveira Silva, Adriane Leites Strothmann, Henrique Gonçalves Pegoraro, Fernanda Severo Sabedra Sousa, Fabiana Kommling Seixas, Francisco Silvestre Brilhante Bezerra, Tiago Veiras Collares, Cláudio Martin Pereira de Pereira, Andrés Mansilla, Sibele Borsuk","doi":"10.1007/s10529-025-03622-3","DOIUrl":"https://doi.org/10.1007/s10529-025-03622-3","url":null,"abstract":"Recently, new immunomodulatory compounds have benn sought as effective adjuvants in vaccine development. In this context, the bioactive substances from macroalgae stand out, as they can satisfactorily activate an immune response against infectious diseases, such as caseous lymphadenitis (CLA), caused by Corynebacterium pseudotuberculosis. This study aimed to evaluate the adjuvant activity of Iridea cordata and Sacorpletis skotibergii lipid extracts associated with the protein rCP01850 against infection by C. pseudotuberculosis in a murine model. Five groups of ten BALB/c mice each were inoculated with 0.9% saline (G1), rCP01850 (G2), rCP01850 + saponin (G3), rCP01850 + Iridea cordata (G4), and rCP01850 + Sacorpletis skotibergii (G5). Two doses of vaccine were administered with a 21-day interval between doses. After that, the animals were challenged with 2 × 104 UFC of the MIC-6 strain. Experimental groups G4 and G5 presented protection rates of 60 and 70%, respectively. The production levels of total IgG and its IgG1 and IgG2a isotypes were significantly increased in G4 and G5 after the forty-second day of immunization. In addition, the expression of the cytokines IL-4, IL-12, IL-10, and IFN-γ significantly increased in G4 and G5 when compared to the negative control (G1). In turn, IL-17 and TNF-α had significant expression levels in G4 when compared to the other experimental groups (p < 0.05). The results show that subantarctic macroalgae extracts associated with rCP01850 induced substantial levels of humoral and cellular immune response and protected immunized animals against the challenge.","PeriodicalId":8929,"journal":{"name":"Biotechnology Letters","volume":"47 4","pages":"82"},"PeriodicalIF":2.0,"publicationDate":"2025-07-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144697475","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Application of aurintricarboxylic acid to boost CHO cell performance. 金三羧酸提高CHO细胞性能的应用。

IF 2 4区生物学

Biotechnology Letters Pub Date : 2025-07-23 DOI: 10.1007/s10529-025-03609-0

Chao Tao, Jizhou Meng, Dan Wen, Jia Dai, Youyou Yu, Yu Wang, Xiangju Wei, Qing Zhao, Ruiqiang Sun, Hang Zhou

{"title":"Application of aurintricarboxylic acid to boost CHO cell performance.","authors":"Chao Tao, Jizhou Meng, Dan Wen, Jia Dai, Youyou Yu, Yu Wang, Xiangju Wei, Qing Zhao, Ruiqiang Sun, Hang Zhou","doi":"10.1007/s10529-025-03609-0","DOIUrl":"https://doi.org/10.1007/s10529-025-03609-0","url":null,"abstract":"Purpose: Currently, a robust process using healthy Chinese hamster ovary (CHO) cells, the dominating host for biopharmaceutical protein therapeutics, is crucial for high productivity and successful scaling-up. However, issues which are originated from cell performance and detrimental to protein products such as dramatic viability drop and accumulation of toxic metabolites still exist. Aurintricarboxylic acid (ATA), reported as an antioxidant chelator to prevent apoptosis and boost cell growth, was applied as an additive in this study to address the above problems.Methods: Two CHO cells were cultivated by fed-batch culture with ATA of different concentrations and adding strategies supplemented. The reactive oxygen species (ROS) test and confocal microscopy have been used to illustrate the working mechanism of ATA.Results: ATA was proved to be effective in maintaining viability and improving lactate performance in the late fed-batch culture stage for both tested clones. Briefly, the harvest viability was increased by at least 10% after ATA was introduced, and the suppression of lactate accumulation was observed with ATA addition. Moreover, besides adding ATA in the fed-batch stage, a novel and favorable process to involve ATA in seed train was developed, which improved the performance of seed and further benefitted the cell performance and productivity during fed-batch culture. And the ATA was detected to penetrate into CHO cells and suppress the ROS generation intracellularly.Conclusion: Overall, this study advances current knowledge with respect to ATA's capability of enhancing cell performances for CHO cell manufacturing and introduces a novel process of hyper seed train.","PeriodicalId":8929,"journal":{"name":"Biotechnology Letters","volume":"47 4","pages":"81"},"PeriodicalIF":2.0,"publicationDate":"2025-07-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144688791","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Bacillus alcalophilus RecJ (BaRecJ) can drive the adaptive evolution of S. cerevisiae. 嗜钙芽孢杆菌RecJ （BaRecJ）可以驱动酿酒酵母的适应性进化。

IF 2 4区生物学

Biotechnology Letters Pub Date : 2025-07-22 DOI: 10.1007/s10529-024-03552-6

Jixiang Shang, Yanchao Zhang, Zongjun Xu, Shouqing Zhang, Zhongtao Sun, Minggang Zheng

引用次数: 0

Optimization and characterization of collagenase KU665299 and its application in effective in-vitro clot digestion. 胶原酶KU665299的优化、表征及其在体外凝块消化中的应用

IF 2 4区生物学

Biotechnology Letters Pub Date : 2025-07-22 DOI: 10.1007/s10529-025-03615-2

Shikha Chauhan, Kriti Kanwar, Deepika Sharma, Harjodh Singh, Deepak Sharma, Vishal Ahuja, Wamik Azmi

{"title":"Optimization and characterization of collagenase KU665299 and its application in effective in-vitro clot digestion.","authors":"Shikha Chauhan, Kriti Kanwar, Deepika Sharma, Harjodh Singh, Deepak Sharma, Vishal Ahuja, Wamik Azmi","doi":"10.1007/s10529-025-03615-2","DOIUrl":"https://doi.org/10.1007/s10529-025-03615-2","url":null,"abstract":"Objective: The study employed response surface methodology (RSM) to optimize physicochemical variables for extracellular collagenase production by gram negative bacterial strain Chryseobacterium contaminans KU665299 under submerged fermentation. It is also revealing the ability of collagenase to degrade collagen, main structural protein in human blood.Result: The study successfully enhanced collagenase activity by 1.2 folds through Response Surface Methodology (RSM) and 5.33 folds through purification of enzyme using ammonium sulfate precipitation and DEAE-Sepharose chromatography (specific activity with 538.0 U/mg). SDS-PAGE analysis identified its molecular weight as 32 kDa. Optimal conditions for the enzyme's activity were pH 7.5 and 40 °C. Kinetic studies of collagenase KU665299 revealed specificity for collagen, with Km and Vmax values of 0.059 mg/l and 588.24 µmol/min/mg, respectively. Zinc and calcium ions enhanced activity, while EDTA and DTT strongly inhibited it. The purified collagenase demonstrated remarkable efficiency in digesting blood clots, fully dissolving 1 ml clots within 40 min at 37 °C, showcasing significant thrombolytic potential.Conclusion: The study successfully optimized and characterized a novel collagenase from C. contaminans KU665299, revealing its high specificity, stability, and efficiency in degrading collagen and its promising ability to rapidly digest blood clots for potential thrombolytic properties.","PeriodicalId":8929,"journal":{"name":"Biotechnology Letters","volume":"47 4","pages":"79"},"PeriodicalIF":2.0,"publicationDate":"2025-07-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144688793","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

A single-vector CRISPR/Cas9 system for genome editing and heterologous enzyme secretion in Saccharomyces cerevisiae: a case study on pectate lyase for coffee mucilage removal. 酿酒酵母菌基因组编辑和异源酶分泌的单载体CRISPR/Cas9系统——以果胶裂解酶去除咖啡粘液为例

IF 2 4区生物学

Biotechnology Letters Pub Date : 2025-07-19 DOI: 10.1007/s10529-025-03621-4

La Ho Truc Lam, Nguyen Huynh Ha Nhi, Vo Thi Hoang Lan, Nguyen Van Hau, Nguyen Hieu Nghia

{"title":"A single-vector CRISPR/Cas9 system for genome editing and heterologous enzyme secretion in Saccharomyces cerevisiae: a case study on pectate lyase for coffee mucilage removal.","authors":"La Ho Truc Lam, Nguyen Huynh Ha Nhi, Vo Thi Hoang Lan, Nguyen Van Hau, Nguyen Hieu Nghia","doi":"10.1007/s10529-025-03621-4","DOIUrl":"https://doi.org/10.1007/s10529-025-03621-4","url":null,"abstract":"The CRISPR/Cas9 system facilitates precise genome editing in various organisms. In this study, a single-vector CRISPR/Cas9 system was developed for Saccharomyces cerevisiae, employing a type II Cas9 enzyme from Streptococcus pyogenes and a single-guide RNA cassette targeting CAN1.Y locus on chromosome V. This system is broadly applicable across yeast strains, as it utilizes G418 selection, eliminating the need for auxotrophic markers. The efficiency of the CRISPR/Cas9 system was demonstrated, with editing efficiencies ranging from 70 to 100%. This system was utilized to integrate a cassette encoding secretory pectate lyase (PL) from Bacillus subtilis 168 into the yeast genome. The engineered S. cerevisiae strain secreted active PL, which exhibited pectin-degrading activity characterized by significant reductions in residual pectin and increased production of reducing sugars. Since pectin constitutes a major component of coffee mucilage, the secreted PL was applied to coffee beans for mucilage removal. The treated beans presented noticeably reduced residual mucilage, a purer green color, and decreased viscosity. These findings suggest the potential of the engineered S. cerevisiae strain for applications in coffee processing, particularly in efficient mucilage removal.","PeriodicalId":8929,"journal":{"name":"Biotechnology Letters","volume":"47 4","pages":"78"},"PeriodicalIF":2.0,"publicationDate":"2025-07-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144666999","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

A novel method for generating baculovirus bacmids using EGFP-mediated purification and linearization. 利用egfp介导的纯化和线性化生成杆状病毒毒株的新方法。

IF 2 4区生物学

Biotechnology Letters Pub Date : 2025-07-17 DOI: 10.1007/s10529-025-03619-y

Wujie Su, Haoyi Gu, Xiaoxia Zhang, Wenbing Wang, Fanchi Li, Bing Li

{"title":"A novel method for generating baculovirus bacmids using EGFP-mediated purification and linearization.","authors":"Wujie Su, Haoyi Gu, Xiaoxia Zhang, Wenbing Wang, Fanchi Li, Bing Li","doi":"10.1007/s10529-025-03619-y","DOIUrl":"https://doi.org/10.1007/s10529-025-03619-y","url":null,"abstract":"Baculovirus bacmids have been widely used in over-expression and gene deletion. Traditionally, baculovirus bacmids are developed by inserting an 8.6 kbp bacterial DNA cassette into baculovirus genomes either through homologous recombination in cultured cells or via in vitro cloning. In this study, by introducing Bsu36i-attached egfp to the 8.6 kbp bacterial DNA cassette, we develop a novel method for generating baculovirus bacmids. An 11.6 kbp bacterial DNA cassette containing the introduced egfp was used to generate an intermediate bacmid. With the EGFP reporter, purification was performed in cultured cells, increasing the proportions of recombinants. The intermediate bacmid containing the 11.6 kbp bacterial DNA cassette was obtained by transforming DH10B competent cells with viral DNA after 3 rounds of purification. The intermediate bacmid DNA was linearized by digestion with Bsu36i and then was co-transfected with the PCR-amplified 8.6 kbp bacterial cassette into BmN cells, where homologous recombination occurred between them. The final BmNPV bacmid was obtained by transforming DH10B competent cells with viral DNA. Capable of increasing the proportions of recombinants via purification and linearization, this method has great potential to be used for bacmid generation for baculoviruses, especially those that are not capable of producing high titers of viruses.","PeriodicalId":8929,"journal":{"name":"Biotechnology Letters","volume":"47 4","pages":"77"},"PeriodicalIF":2.0,"publicationDate":"2025-07-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144648424","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

A novel P450 enzyme assay utilizing an NADP⁺-based biosensor. 利用NADP+为基础的生物传感器的新型P450酶测定。

IF 2 4区生物学

Biotechnology Letters Pub Date : 2025-07-16 DOI: 10.1007/s10529-025-03599-z

Sifan Shangguan, Taichang Wang, Di Zhao, Guobin Zhang, Yisang Zhang, Ruiming Wang, Junqing Wang, Jing Su

{"title":"A novel P450 enzyme assay utilizing an NADP+-based biosensor.","authors":"Sifan Shangguan, Taichang Wang, Di Zhao, Guobin Zhang, Yisang Zhang, Ruiming Wang, Junqing Wang, Jing Su","doi":"10.1007/s10529-025-03599-z","DOIUrl":"https://doi.org/10.1007/s10529-025-03599-z","url":null,"abstract":"Purpose: High-throughput screening methods for cytochrome P450 enzymes (P450s), such as colorimetric, mass spectrometric, and fluorescence-based assays, often face limitations in throughput, real-time monitoring, and versatility.Methods: To address these challenges, we developed a novel biosensor leveraging glucose-6-phosphate dehydrogenase and Bimolecular Fluorescence Complementation for real-time monitoring of intracellular NADP+ levels, enabling P450 activity detection. The sensor was applied to monitor P450 activity by tracking intracellular NADP+ dynamics, as P450s catalyze diverse substrate reactions and convert NADPH to NADP+ via their electron transport system. To enhance detection precision, intracellular NADP+ synthesis was reduced by knocking down NADPH-dependent aldehyde reductase (YqhD), minimizing background fluorescence interference.Results: The sensor exhibited a linear NADP+ detection range of 1 μM to 10 mM, suitable for P450 assays. The sensor's performance was validated by comparing P450 activities in engineered strains with traditional gas chromatography.Conclusion: The developed biosensor demonstrates its potential as a robust, real-time screening tool for P450 enzyme studies.","PeriodicalId":8929,"journal":{"name":"Biotechnology Letters","volume":"47 4","pages":"76"},"PeriodicalIF":2.0,"publicationDate":"2025-07-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144641711","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0