Biotechnology Journal最新文献

{"title":"Heterologous Biosynthesis of Terpenoids in Saccharomyces cerevisiae","authors":"Junyang Wang,&nbsp;Xu Ji,&nbsp;Renhe Yi,&nbsp;Dengbin Li,&nbsp;Xiaolin Shen,&nbsp;Zihe Liu,&nbsp;Yaying Xia,&nbsp;Shuobo Shi","doi":"10.1002/biot.202400712","DOIUrl":"10.1002/biot.202400712","url":null,"abstract":"<div>\u0000 \u0000 <p>Terpenoids are widely distributed in nature and have various applications in healthcare products, pharmaceuticals, and fragrances. Despite the significant potential that terpenoids possess, traditional production methods, such as plant extraction and chemical synthesis, face challenges in meeting current market demand. With the advancement of synthetic biology and metabolic engineering, it becomes feasible to construct efficient microbial cell factories for large-scale production of terpenoids. This article primarily centers on the heterologous expression of terpenoids in <i>Saccharomyces cerevisiae</i>, detailing the expression of terpenoid biosynthesis pathways through the utilization of cellular microcompartments, strategies for the efficient expression of key P450 enzymes in the synthesis pathway, and the regulation and optimization of host metabolism to enhance flux to terpenoids synthesis. Additionally, we analyze current challenges and propose solutions to further refine yeast chassis for more effective terpenoids production.</p>\u0000 </div>","PeriodicalId":134,"journal":{"name":"Biotechnology Journal","volume":"20 1","pages":""},"PeriodicalIF":3.2,"publicationDate":"2025-01-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142996803","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}

{"title":"Tobacco: A Favorite for Low-Carbon Biorefinery","authors":"Deshui Liu,&nbsp;Xiang Li,&nbsp;Zhonghao Li","doi":"10.1002/biot.202400677","DOIUrl":"10.1002/biot.202400677","url":null,"abstract":"&lt;p&gt;Biomass is increasingly recognized as a renewable source for the replacement of fossil fuel, as well as for the production of fuel and chemical production, due to its organic nature, abundant supply, and potential for negative emission. However, a cost-efficient bioconversion process, along with promising alternatives and streamlined utilization routes to chemicals, remains essential for the commercial deployment of biomass. Tobacco, a controversially planted crop primarily used as raw materials in the cigarette industry, has gained attention in light of the global adoption of the WHO Framework Convention on Tobacco Control (WHO FCTC) and the rising interest in its seed oil. Its high-yielding, broad cultivation, and suitability for genetic engineering have led to its consideration as a candidate for biofuels production. Nevertheless, neither the tobacco plant for production of seed oil or the hydrothermal process applied to tobacco biomass has established it as a viable feedstock candidate. A recent study published in &lt;i&gt;The Innovation&lt;/i&gt; by Wang et al. [&lt;span&gt;1&lt;/span&gt;] has proposed a novel and simplest strategy that promotes tobacco as a highly promising energy crop for bioproducts production, with the potential to significantly reduce greenhouse gas emissions (Figure 1).&lt;/p&gt;&lt;p&gt;Unlike traditional biomass feedstocks, tobacco was characterized by its high content of water-soluble carbohydrates (65%) (Figure 1a) and nitrogen, along with a low level of lignocellulose, and it is capable of growing on marginal lands, rendering it an ideal material for low-energy and low-carbon bioconversion processes. By simply autoclaving tobacco leaves in water, a nutrient medium was obtained that effectively supported the growth of microorganisms and the production of bioproducts (Figure 1c), eliminating the need for pretreatment and hydrolysis of the feedstock or the addition of supplements to medium. Additionally, the study employed a life cycle assessment (LCA) approach to evaluate the carbon-negative effects of tobacco biomass in the bioethanol production process. The findings indicated that tobacco bioethanol could reduce carbon emissions by up to approximately 76% and lower energy consumption by approximately 81% compared to traditional corn stover bioethanol during biorefinery processes (Figure 1b).&lt;/p&gt;&lt;p&gt;Massive biomass production and growth on marginal lands are two scientifically significant criteria for identifying a paradigmatic energy crop. Tobacco (&lt;i&gt;Nicotiana tabacum&lt;/i&gt;) is among the most widely cultivated non-food crops globally, grown in over 100 countries. Tobacco cultivation can yield multiple harvests per year and produces a substantial biomass (Figure 1d), potentially reaching up to 170 tons/ha when cultivated for biofuel and biochemical production [&lt;span&gt;2&lt;/span&gt;]. Its whole genome was published in 2014 [&lt;span&gt;3&lt;/span&gt;], and numerous advanced genetic engineering tools are now available for its manipulation. Consequently, tobacco is amena","PeriodicalId":134,"journal":{"name":"Biotechnology Journal","volume":"20 1","pages":""},"PeriodicalIF":3.2,"publicationDate":"2025-01-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/biot.202400677","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142996815","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Subtractive Inhibition Assay Based on PagN-Specific Monoclonal Antibody for the Detection of Salmonella Using Surface Plasmon Resonance","authors":"Maozhi Hu,&nbsp;Hongji Zhu,&nbsp;Chuang Meng,&nbsp;Ruiqing Ding,&nbsp;Qiuxiang Yan,&nbsp;Hongyan Zhao,&nbsp;Xilong Kang,&nbsp;Dan Gu,&nbsp;Zhiming Pan,&nbsp;Xinan Jiao","doi":"10.1002/biot.202400616","DOIUrl":"10.1002/biot.202400616","url":null,"abstract":"<p><i>Salmonella</i> is a common foodborne zoonotic pathogen that poses a great threat to human health and breeding industry. The rapid detection of <i>Salmonella</i> is necessary for early prevention and control. In this study, a subtractive inhibition assay (SIA) based on surface plasmon resonance (SPR) for the rapid detection of <i>Salmonella</i> was developed. Mouse-specific monoclonal antibody 3B3 against <i>Salmonella</i> membrane protein PagN was first incubated with <i>Salmonella</i>. <i>The</i> unbound free antibody was separated using a sequential process of centrifugation and then detected using an immobilized goat anti-mouse immunoglobulin G polyclonal antibody on the SPR sensor chip. This SIA-SPR method showed excellent sensitivity for <i>Salmonella</i> with a limit of detection of about 300 CFU mL⁻¹. This method is sensitive to different serotypes of <i>Salmonella</i> strains but not for non-<i>Salmonella</i> strains. It was able to detect <i>Salmonella</i> in the contaminated water and milk powder at less than 10² and 10³ CFU mL⁻¹, respectively, which was consistent with the bacterial plate count results. In addition, this method could be used to evaluate the lysis effect of phages on bacteria. Since the culturing detection method needs more than 48 h, this method has the potential for the rapid and sensitive clinical detection of <i>Salmonella</i>. For our knowledge, this is the first report for <i>Salmonella</i> detection using SIA-SPR method.</p>","PeriodicalId":134,"journal":{"name":"Biotechnology Journal","volume":"20 1","pages":""},"PeriodicalIF":3.2,"publicationDate":"2025-01-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142996814","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}

{"title":"Triune Engineering Approach for (+)-valencene Overproduction in Yarrowia lipolytica","authors":"Ying Chen,&nbsp;Liqiu Su,&nbsp;Qi Liu,&nbsp;Ge Zhang,&nbsp;Hongyang Chen,&nbsp;Qinhong Wang,&nbsp;Kaizhi Jia,&nbsp;Zongjie Dai","doi":"10.1002/biot.202400669","DOIUrl":"10.1002/biot.202400669","url":null,"abstract":"<div>\u0000 \u0000 <p>The sesquiterpene (+)-valencene, with its flavor and diverse biological functions, holds promise for applications in the food, fragrance, and pharmaceutical industries. However, the low concentration in nature and high cost of extraction limit its application. This study aimed to construct a microbial cell factory to efficiently produce (+)-valencene. The strain <i>Yarrowia lipolytica</i> YL238, possessing a stronger capacity for (+)-valencene synthesis, was selected and utilized as the chassis for further modifications. By fine-tuning the mevalonate and squalene synthesis pathways we achieved a remarkable 13.2-fold increase in (+)-valencene titer compared to the original strain. Following directed evolution was employed to screen for efficient (+)-valencene synthase, which further enhanced (+)-valencene production by 138%. Consequently, the engineered strain overproduced 813 mg/L of (+)-valencene in shake flasks, marking the highest titer reported in microbials to date. Furthermore, in fed-batch fermentation, this engineered strain showed the capacity to produce 3.3 g/L of (+)-valencene. This study offers a successful model for the application of the “strain-pathway-enzyme” triune strategy in the metabolic engineering of <i>Y. lipolytica</i>, and these methodologies could be broadly utilized for the synthesis of other natural terpenes.</p>\u0000 </div>","PeriodicalId":134,"journal":{"name":"Biotechnology Journal","volume":"20 1","pages":""},"PeriodicalIF":3.2,"publicationDate":"2025-01-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142996819","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}

{"title":"Towards a Platform Chromatography Purification Process for Adeno-Associated Virus (AAV)","authors":"Alexandra Bogdanovic,&nbsp;Nicholas Donohue,&nbsp;Brian Glennon,&nbsp;Susan McDonnell,&nbsp;Jessica Whelan","doi":"10.1002/biot.202400526","DOIUrl":"10.1002/biot.202400526","url":null,"abstract":"<div>\u0000 \u0000 <p>Adeno-associated virus (AAV) is a versatile viral vector technology that can be engineered for specific functionality in vaccine and gene therapy applications. One of the major challenges in AAV production is the need for a GMP-ready platform-based approach to downstream processing, as this would lead to a standardized method for multiple products. Chromatography has huge potential in AAV purification, as it is a scalable method that would enable manufacturing to a high degree of purity, potency, and consistency. Multiple factors need to be considered when developing a chromatography platform, including the chromatography type, format, and mode of operation, along with other commercial considerations that have not been comprehensively reviewed until now. In addition to chromatography factors, this review will also consider the current understanding of AAV characteristics: this will include net surface charge, structural properties, and size, as well as their interactions with metal ions or receptors, and how this impacts the development of a chromatography platform.</p>\u0000 </div>","PeriodicalId":134,"journal":{"name":"Biotechnology Journal","volume":"20 1","pages":""},"PeriodicalIF":3.2,"publicationDate":"2025-01-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142996818","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}

{"title":"Rational Design of a Novel DNA Polymerase From Clostridium thermocellum to Improve LAMP Detection Performance","authors":"Cheng Wang,&nbsp;Bin Hong,&nbsp;Yanmei Li,&nbsp;Yi Ma,&nbsp;Wei Xu,&nbsp;Jufang Wang","doi":"10.1002/biot.202400559","DOIUrl":"10.1002/biot.202400559","url":null,"abstract":"<div>\u0000 \u0000 <p>Loop-mediated isothermal amplification (LAMP) is a detection method widely used in pathogen detection and clinical diagnosis. Nevertheless, it is highly constrained by thermal stability, catalytic activity, and resistance to inhibitors of Bst DNA polymerase. In this study, a novel DNA polymerase was characterized from <i>Clostridium thermocellum</i>, exhibiting potential in LAMP detection. Through bioinformatics analysis, the enzyme and the DNA-binding domain (DBD) from <i>Pyrococcus abyssi</i> were mutated for enhanced interaction between proteins and DNA. A chimeric mutant DBD_E146K-S738R reaches the detection threshold 13 min earlier than wild-type Cth DNA polymerase in real-time LAMP detection with a template concentration of 1.58 × 10⁵ fg/µL. It also showed the highest enzymatic activity at pH 9.0 and 65°C. The chimeric enzyme DBD_E146K-S738R exhibits good thermal stability, capable of performing LAMP reactions after treatment at 73°C or 70°C for 8 h. Moreover, it maintains high activity even under the inhibitory conditions of 50 U/mL heparin, 1.6 mM EDTA, 200 mM NaCl, 10% ethanol, 1.2 M urea, or 0.8% phenol. Notably, it was able to detect 1.58 × 10² ag/µL of the genome and 1.03 CFU/mL of the colony in <i>Salmonella typhimurium</i> detection. The enzyme's performance is superior to commercial Bst 2.0 and comparable to commercial Bst 3.0. The results suggest that DBD_E146K-S738R in LAMP exhibits great potential for molecular biological studies and clinical diagnostic analysis.</p>\u0000 </div>","PeriodicalId":134,"journal":{"name":"Biotechnology Journal","volume":"20 1","pages":""},"PeriodicalIF":3.2,"publicationDate":"2025-01-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142941777","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}

{"title":"Virus Filtration Development for Adeno-Associated Virus-Based Gene Therapy Products","authors":"FNU Namila,&nbsp;Tianyi Zhou,&nbsp;Lu Wang,&nbsp;Mi Jin","doi":"10.1002/biot.202400636","DOIUrl":"10.1002/biot.202400636","url":null,"abstract":"<div>\u0000 \u0000 <p>Adeno-associated virus (AAV) vectors have become a leading platform for gene delivery. A major portion of gene therapy currently in clinical trials are AAV-based for a wide range of diseases. A commonly used method for AAV production is by mammalian or insect cell culture, with or without added viruses to introduce needed genetic elements for AAV production. There are potential risks of virus contamination from the production cell line, process residuals, or adventitious contamination in the production of biotherapeutics, including AAV-based gene therapy products; therefore, it is imperative to demonstrate that the drug substance manufacturing process has sufficient capability to clear process-related or adventitious viruses. In the AAV-based gene therapy manufacturing process, cell lysis, affinity chromatography, and ion exchange chromatography steps are often effective to inactivate or remove viruses. To increase the viral clearance robustness, virus filtration (VF) is increasingly recommended by regulatory agencies for gene therapy products as a dedicated viral clearance step in the downstream purification process. In the current study, two commercially available virus filters were evaluated in the context of AAV manufacturing. The filter throughput and process yield were assessed under different operational modes. Virus clearance performance was evaluated by spiking in Adenovirus type 5 (Adv-5) and Simian virus 40 (SV-40). The viral filters assessed in this study demonstrated manufacturable throughputs, acceptable process yields, and robust virus clearance capabilities for viruses greater than 40 nm.</p>\u0000 </div>","PeriodicalId":134,"journal":{"name":"Biotechnology Journal","volume":"20 1","pages":""},"PeriodicalIF":3.2,"publicationDate":"2025-01-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142941779","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}

{"title":"Issue Information: Biotechnology Journal 1/2025","authors":"","doi":"10.1002/biot.202570001","DOIUrl":"https://doi.org/10.1002/biot.202570001","url":null,"abstract":"","PeriodicalId":134,"journal":{"name":"Biotechnology Journal","volume":"20 1","pages":""},"PeriodicalIF":3.2,"publicationDate":"2025-01-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://onlinelibrary.wiley.com/doi/epdf/10.1002/biot.202570001","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143113086","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Multienzyme Cascade Synthesis of Rare Sugars From Glycerol in Bacillus subtilis","authors":"Kangqing Fei,&nbsp;Liqun Shen,&nbsp;Xiao-Dong Gao,&nbsp;Hideki Nakanishi,&nbsp;Zijie Li","doi":"10.1002/biot.202400539","DOIUrl":"10.1002/biot.202400539","url":null,"abstract":"<div>\u0000 \u0000 \u0000 <section>\u0000 \u0000 <h3> Background</h3>\u0000 \u0000 <p>Rare sugars are valuable and unique monosaccharides extensively utilized in the food, cosmetics, and pharmaceutical industries. Considering the high purification costs and the complex processes of enzymatic synthesis, whole-cell conversion has emerged as a significantly important alternative. The <i>Escherichia coli</i> strain was initially used in whole-cell synthesis of rare sugars. However, its pathogenic nature poses limitations to its widespread applications. Consequently, there is an urgent need to explore biologically safe strains for the efficient production of rare sugars.</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Results</h3>\u0000 \u0000 <p>In this study, the generally regarded as safe (GRAS) strain <i>Bacillus subtilis</i> was employed as the chassis cells to produce rare sugars via whole-cell conversion. Three genes encoding alditol oxidase (AldO), L-rhamnulose-1-phosphate aldolase (RhaD), and fructose-1-phosphatase (YqaB) involved in rare sugars biosynthesis were heterogeneously expressed in <i>B. subtilis</i> to convert the only substrate glycerol into rare sugars. To enhance the expression levels of the relevant enzymes in <i>B. subtilis</i>, different promoters for <i>aldO</i>, <i>rhaD</i>, and <i>yqaB</i> were investigated and optimized in this system. Under the optimized reaction conditions, the maximum total production titer was 16.96 g/L of D-allulose and D-sorbose with a conversion yield of 33.9% from glycerol. Furthermore, the engineered strain produced 26.68 g/L of D-allulose and D-sorbose through fed-batch for the whole-cell conversion, representing the highest titer from glycerol reported to date.</p>\u0000 </section>\u0000 \u0000 <section>\u0000 \u0000 <h3> Conclusion</h3>\u0000 \u0000 <p>This study demonstrated an efficient and cost-effective method for the synthesis of rare sugars, providing a food-grade platform with the potential to meet the growing demand for rare sugars in industries.</p>\u0000 </section>\u0000 </div>","PeriodicalId":134,"journal":{"name":"Biotechnology Journal","volume":"19 12","pages":""},"PeriodicalIF":3.2,"publicationDate":"2024-12-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142890700","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}

{"title":"Blue Light-Induced, Dosed Protein Expression of Active BDNF in Human Cells Using the Optogenetic CRY2/CIB System","authors":"Sina Christoffers,&nbsp;Nina Wichert,&nbsp;Elena Wiebe,&nbsp;Maria Leilani Torres-Mapa,&nbsp;Madeleine Goblet,&nbsp;Jennifer Harre,&nbsp;Odett Kaiser,&nbsp;Marc-Nils Wahalla,&nbsp;Holger Blume,&nbsp;Alexander Heisterkamp,&nbsp;Athanasia Warnecke,&nbsp;Cornelia Blume","doi":"10.1002/biot.202400384","DOIUrl":"10.1002/biot.202400384","url":null,"abstract":"<p>The use of optogenetic tools offers an excellent method for spatially and temporally regulated gene and protein expression in cell therapeutic approaches. This could be useful as a concomitant therapeutic measure, especially in small body compartments such as the inner ear, for example, during cochlea implantation, to enhance neuronal cell survival and function. Here, we used the blue light activatable CRY2/CIB system to induce transcription of brain-derived neurotrophic factor (BDNF) in human cells. Transfection with three plasmids, encoding for the optogenetic system and the target, as well as illumination protocols were optimized with luciferase as a reporter to achieve the highest protein expression in human embryonic kidney cells 293. Illumination was performed either with a light-emitting diode or with a scanning laser setup. The optimized protocols were applied for the production of BDNF. We could demonstrate a 64.7-fold increase of BNDF expression upon light induction compared to the basal level. Light-induced BDNF was biologically active and enhanced survival and neurite growth of spiral ganglion neurons. The optogenetic approach can be transferred to autologous cell systems, such as bone marrow-derived mesenchymal stem cells, and thus represents the first optogenetic neurotrophic therapy for the inner ear.</p>","PeriodicalId":134,"journal":{"name":"Biotechnology Journal","volume":"19 12","pages":""},"PeriodicalIF":3.2,"publicationDate":"2024-12-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11671672/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142890625","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}