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Creating Better Brewing Yeast With the 1011 Yeast Genomes Data Sets. 使用1011酵母基因组数据集创建更好的酿造酵母。
IF 2.2 4区 生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2025-01-01 Epub Date: 2025-02-15 DOI: 10.1002/yea.3990
Kristoffer Krogerus, Nils Rettberg

Yeast strain development has been essential for improving efficiency, flavour diversity, and quality of beer fermentation. Such efforts often rely on laborious in vitro screening experiments. However, with the increasing availability of large-scale 'omics' data sets, it may be possible to replace or complement such experiments with in silico screening. Compared to more traditional in vitro screening, this has several benefits, including lower costs, more rapid results and possibility to include more strains. Here, we briefly review the genetics associated with various desirable and undesirable traits in brewing yeast, and demonstrate how recent genomics, transcriptomics, and proteomics data sets derived from the 1011 yeast genomes project can be exploited for identifying strains with potentially desirable phenotypes. The discussed phenotypes are related to fermentation performance, formation of desirable flavours, and mitigation of off-flavours. Finally, we perform wort fermentations with five strains from diverse backgrounds, with diverse predicted phenotypes, to validate the in silico predictions. Most predicted phenotypes correlated well with the measured phenotypes, including formation of desirable compounds like isoamyl acetate and ethyl octanoate, as well as formation of undesirable compounds like 4-vinyl guaiacol, diacetyl, and ethanethiol. Together, the results indicate that utilising large 'omics' data sets can be a very useful tool for both strain selection and development for beer fermentation, and naturally other food and beverage fermentations as well. We hope this can inspire and yield improved and more diverse brewing strains to the industry.

酵母菌株的开发对于提高啤酒发酵的效率、风味多样性和质量至关重要。这些工作通常依赖于费力的体外筛选实验。然而,随着大规模 "omics "数据集的日益普及,我们有可能用硅学筛选来取代或补充这些实验。与更传统的体外筛选相比,这有几个好处,包括成本更低、结果更快,而且可以纳入更多菌株。在此,我们简要回顾了与酿酒酵母各种理想和不理想性状相关的遗传学,并展示了如何利用最近从 1011 酵母基因组项目中获得的基因组学、转录组学和蛋白质组学数据集来鉴定具有潜在理想表型的菌株。所讨论的表型与发酵性能、理想风味的形成和异味的减轻有关。最后,我们用五种来自不同背景、具有不同预测表型的菌株进行麦汁发酵,以验证硅学预测。大多数预测的表型与测量的表型有很好的相关性,包括乙酸异戊酯和辛酸乙酯等理想化合物的形成,以及 4-乙烯基愈创木酚、双乙酰基和乙硫醇等不良化合物的形成。总之,这些结果表明,利用大型 "全息 "数据集是一种非常有用的工具,既可用于啤酒发酵的菌种选择和开发,也可用于其他食品和饮料发酵。我们希望这能为酿酒业带来启发,使酿酒菌株得到改良并更加多样化。
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引用次数: 0
The Hidden Global Diversity of the Yeast Genus Carlosrosaea: A Biodiversity Databases Perspective. 酵母属Carlosrosaea隐藏的全球多样性:生物多样性数据库视角。
IF 2.2 4区 生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-11-01 Epub Date: 2024-12-02 DOI: 10.1002/yea.3986
Ciro Ramon Félix, Hector M C Navarro, Melissa Fontes Landell

Biodiversity gaps in microorganisms, such as yeasts, blur our understanding of microbial diversity, introducing biases in their biogeography, ecology, and taxonomy. The genus Carlosrosaea is a potential plant growth booster, yet it is still a little-known yeast group. Considering that databases like GBIF and GenBank are powerful tools for exploring biodiversity data, we aimed to map the geographic distribution, ecological patterns, and taxonomic potential of the genus Carlosrosaea. We found 176 records of the genus, with about 70% associated with plant material, mostly leaves. Furthermore, 55% of the records pertained to the tropical region and only 12% to the temperate. The data indicates the existence of more than a dozen possible new species of the genus, cataloged yet undescribed. This study advances our understanding of the geographic, ecological, and taxonomic aspects of Carlosrosaea. It also highlights how public databases and literature reviews provide accessible ways to analyze information about microbial groups with limited data.

微生物(如酵母)的生物多样性差距模糊了我们对微生物多样性的理解,在其生物地理学、生态学和分类学上引入了偏见。Carlosrosaea属是一种潜在的植物生长助推器,但它仍然是一个鲜为人知的酵母群。考虑到GBIF和GenBank等数据库是探索生物多样性数据的有力工具,我们旨在绘制卡罗斯海属的地理分布、生态格局和分类潜力。我们发现了176份记录,其中约70%与植物材料有关,大部分与叶子有关。此外,55%的记录属于热带地区,只有12%属于温带地区。数据表明,该属存在十多种可能的新种,已编目但未描述。本研究促进了我们对紫菜属植物的地理、生态和分类的认识。它还强调了公共数据库和文献综述如何提供可访问的方法来分析有限数据下的微生物群信息。
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引用次数: 0
CAYSS: Package for Automatic Cytometry Analysis of Yeast Spore Segregation. CAYSS:酵母孢子分离的自动细胞分析包。
IF 2.2 4区 生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-11-01 Epub Date: 2025-01-22 DOI: 10.1002/yea.3988
Xavier Raffoux, Matthieu Falque

Meiotic recombination is a powerful source of haplotypic diversity, and thus plays an important role in the dynamics of short-term adaptation. However, high-throughput quantitative measurement of recombination parameters is challenging because of the large size of offspring to be genotyped. One of the most efficient approaches for large-scale recombination measurement is to study the segregation of fluorescent markers in gametes. Applying this to yeast spores by flow cytometry has already been proved to be highly efficient, but manual analyses of distributions of signal intensities is time-consuming and produces nonperfectly reproducible results. Such analyses are required to identify events corresponding to spores and to assign each of them to a genotypic class depending on their fluorescence intensity. The CAYSS package automatically reproduces the manual process that we've been developing to analyze yeast recombination for years, including Maximum-Likelihood estimation of fluorescence extinction (Raffoux et al. 2018a). When comparing the results of manual versus CAYSS automatic analyses of the same cytometry data, recombination rates and interference were on average very similar, with less than 3% differences on average and strong correlations (R2 > 0.9). In conclusion, as compared to manual analysis, CAYSS allows to save a lot of human time and produces totally reproducible results.

减数分裂重组是单倍型多样性的重要来源,因此在短期适应动力学中起着重要作用。然而,重组参数的高通量定量测量具有挑战性,因为需要进行基因分型的后代很大。研究配子中荧光标记的分离是大规模重组测量最有效的方法之一。通过流式细胞术将其应用于酵母孢子已经被证明是非常高效的,但是对信号强度分布的人工分析是耗时的,并且产生不完全可重复的结果。需要这样的分析来确定与孢子相对应的事件,并根据它们的荧光强度将每个孢子分配到一个基因型类别。CAYSS软件包自动再现了我们多年来一直在开发的分析酵母重组的手动过程,包括荧光消光的最大似然估计(Raffoux等人,2018a)。当比较相同细胞术数据的人工和CAYSS自动分析结果时,重组率和干扰平均非常相似,平均差异小于3%,相关性强(R2 > 0.9)。总之,与手工分析相比,CAYSS可以节省大量的人力时间,并产生完全可重复的结果。
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引用次数: 0
The Viral K1 Killer Yeast System: Toxicity, Immunity, and Resistance. 病毒K1杀手酵母系统:毒性、免疫和抗性。
IF 2.6 4区 生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-11-01 Epub Date: 2025-01-24 DOI: 10.1002/yea.3987
Angelina Chan, Michelle Hays, Gavin Sherlock

Killer yeasts, such as the K1 killer strain of S. cerevisiae, express a secreted anti-competitive toxin whose production and propagation require the presence of two vertically-transmitted dsRNA viruses. In sensitive cells lacking killer virus infection, toxin binding to the cell wall results in ion pore formation, disruption of osmotic homeostasis, and cell death. However, the exact mechanism(s) of K1 toxin killing activity, how killer yeasts are immune to their own toxin, and which factors could influence adaptation and resistance to K1 toxin within formerly sensitive populations are still unknown. Here, we describe the state of knowledge about K1 killer toxin, including current models of toxin processing and killing activity, and a summary of known modifiers of K1 toxin immunity and resistance. In addition, we discuss two key signaling pathways, HOG (high osmolarity glycerol) and CWI (cell wall integrity), whose involvement in an adaptive response to K1 killer toxin in sensitive cells has been previously documented but requires further study. As both host-virus and sensitive-killer competition have been documented in killer systems like K1, further characterization of K1 killer yeasts may provide a useful model system for study of both intracellular genetic conflict and counter-adaptation between competing sensitive and killer populations.

杀手酵母(如 S. Cerevisiae 的 K1 杀手菌株)表达一种分泌型反竞争毒素,这种毒素的产生和传播需要两种垂直传播的 dsRNA 病毒的存在。在缺乏杀手病毒感染的敏感细胞中,毒素与细胞壁的结合会导致离子孔的形成、渗透平衡的破坏和细胞的死亡。然而,K1 毒素杀伤活性的确切机制、杀手酵母如何对自身毒素产生免疫以及哪些因素会影响以前敏感种群对 K1 毒素的适应性和抵抗力,这些问题仍然不得而知。在此,我们将介绍有关 K1 杀人酵母毒素的知识现状,包括毒素处理和杀伤活性的现有模型,以及 K1 毒素免疫和抗性的已知调节因子摘要。此外,我们还讨论了两个关键的信号通路,即 HOG(高渗透压甘油)和 CWI(细胞壁完整性),这些通路参与敏感细胞对 K1 杀手毒素的适应性反应的情况以前已有记载,但还需要进一步研究。由于在 K1 等杀手系统中已经记录了宿主-病毒和敏感-杀手之间的竞争,对 K1 杀手酵母的进一步鉴定可能会为研究细胞内遗传冲突以及竞争的敏感和杀手种群之间的反适应提供一个有用的模型系统。
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引用次数: 0
Role of Oral Yeast in Replenishing Gastric Mucosa with Yeast and Helicobacter pylori. 口服酵母在补充胃黏膜酵母和幽门螺旋杆菌中的作用
IF 2.2 4区 生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-11-01 Epub Date: 2024-11-15 DOI: 10.1002/yea.3983
Sara Kadkhodaei, Atousa Hatefi, Shahrzad Pedramnia, Elham Godini, Saman Khalili-Samani, Parastoo Saniee, Abdolfattah Sarrafnejad, Ali-Hatef Salmanian, Masoud Sotoudeh, David Y Graham, Reza Malekzadeh, Farideh Siavoshi

The relationship between oral and gastric yeasts and their role in the colonization of Helicobacter pylori in the stomach was studied. Four groups of 221, 7, 44, and 10 patients were used for the isolation of H. pylori and oral and gastric yeasts. In Group 1, gastric biopsies were used for the isolation of H. pylori and yeast, rapid urease test (RUT), staining with Gram's and hematoxylin & eosin (H&E), and immunohistochemistry (IHC) methods. In the other three groups, DNAs extracted from H. pylori and yeasts were used for the amplification of H. pylori-specific genes. Wet mounts of yeasts in Group 2 were examined to observe intracellular bacteria and released EVs. Among 221 patients, 65 (29.3%) had oral yeast, 35 (15.8%) H. pylori, and 31 (14%) gastric yeast. Culture of oral yeasts showed a significant correlation with the detection of H. pylori by IHC (10.3%), Gram stain (9%), RUT (6.3%), H&E (4.9%), and culture (4%) (p < 0.05). Gram-stained biopsies showed the occurrence of yeast and H. pylori, and the release of EVs from yeast. Detection of similar H. pylori genes in oral and gastric yeasts from patients in Group 2 showed their common source. Oral yeasts in Groups 3 and 4 also carried H. pylori genes. Wet mount preparations of yeasts showed intracellular bacteria inside the yeast vacuole and the release of EVs that could carry H. pylori. Oral yeast protects its intracellular H. pylori and releases it inside EVs to safely reach gastric mucosa. Yeast, as the environmental reservoir of H. pylori, plays a crucial role in bacterial reinfection after successful eradication.

研究了口腔酵母菌和胃酵母菌之间的关系及其在幽门螺杆菌胃内定植中的作用。四组分别为 221、7、44 和 10 名患者,用于分离幽门螺杆菌和口腔酵母菌及胃酵母菌。第一组采用胃活检、快速尿素酶试验(RUT)、革兰氏染色、苏木精和伊红(H&E)染色以及免疫组织化学(IHC)方法分离幽门螺杆菌和酵母菌。其他三组则使用从幽门螺杆菌和酵母菌中提取的 DNA 来扩增幽门螺杆菌特异性基因。对第 2 组酵母的湿装片进行检查,以观察胞内细菌和释放的 EVs。在221名患者中,65人(29.3%)有口腔酵母菌,35人(15.8%)有幽门螺杆菌,31人(14%)有胃酵母菌。通过 IHC(10.3%)、革兰氏染色(9%)、RUT(6.3%)、H&E(4.9%)和培养(4%),口腔酵母菌的培养与幽门螺杆菌的检测有显著相关性(p
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引用次数: 0
pSPObooster: A Plasmid System to Improve Sporulation Efficiency of Saccharomyces cerevisiae Lab Strains. pSPObooster:提高酿酒酵母实验室菌株繁殖效率的质粒系统
IF 2.2 4区 生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-10-01 Epub Date: 2024-09-09 DOI: 10.1002/yea.3978
Raphael Loll-Krippleber, Yangyang Kate Jiang, Grant W Brown

Common Saccharomyces cerevisiae lab yeast strains derived from S288C have meiotic defects and therefore are poor sporulators. Here, we developed a plasmid system containing corrected alleles of the MKT1 and RME1 genes to rescue the meiotic defects and show that standard BY4741 and BY4742 strains containing the plasmid display faster and more efficient sporulation. The plasmid, pSPObooster, can be maintained as an episome and easily cured or stably integrated into the genome at a single locus. We demonstrate the use of pSPObooster in low- and high-throughput yeast genetic manipulations and show that it can expedite both procedures without impacting strain behavior.

源自 S288C 的普通酿酒酵母实验菌株存在减数分裂缺陷,因此孢子发育不良。在这里,我们开发了一种含有 MKT1 和 RME1 基因校正等位基因的质粒系统来挽救减数分裂缺陷,并证明含有该质粒的标准 BY4741 和 BY4742 株系能更快更有效地产生孢子。这种名为 pSPObooster 的质粒可以作为外显子保持,并且很容易固化或稳定地整合到基因组的单个位点上。我们展示了 pSPObooster 在低通量和高通量酵母遗传操作中的应用,并证明它能加快这两种操作过程而不影响菌株的行为。
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引用次数: 0
The 5-Fluorouracil RNA Expression Viewer (5-FUR) Facilitates Interpreting the Effects of Drug Treatment and RRP6 Deletion on the Transcriptional Landscape in Yeast. 5-氟尿嘧啶 RNA 表达查看器(5-FUR)有助于解读药物治疗和 RRP6 缺失对酵母转录景观的影响。
IF 2.2 4区 生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-10-01 Epub Date: 2024-09-30 DOI: 10.1002/yea.3982
Ugo Szachnowski, Oliver Sallou, Mateo Boudet, Anthony Bretaudeau, Maxime Wery, Antonin Morillon, Michael Primig

Saccharomyces cerevisiae is an excellent model to study the effect of external cues on cell division and stress response. 5-Fluorocuracil (5-FU) has been used to treat solid tumors since several decades. The drug was initially designed to interfere with DNA replication but was later found to exert its antiproliferative effect also via RNA-dependent processes. Since 5-FU inhibits the activity of the 3'-5'-exoribonuclease Rrp6 in yeast and mammals, earlier work has compared the effect of 5-FU treatment and RRP6 deletion at the transcriptome level in diploid synchronized yeast cells. To facilitate interpreting the expression data we have developed an improved 5-Fluorouracil RNA (5-FUR) expression viewer. Users can access information via genome coordinates and systematic or standard names for mRNAs and Xrn1-dependent-, stable-, cryptic-, and meiotic unannotated transcripts (XUTs, SUTs, CUTs, and MUTs). Normalized log2-transformed or linear data can be displayed as filled diagrams, line graphs or color-coded heatmaps. The expression data are useful for researchers interested in processes such as cell cycle regulation, mitotic repression of meiotic genes, the effect of 5-FU treatment and Rrp6 deficiency on the transcriptome and expression profiles of sense/antisense loci that encode overlapping transcripts. The viewer is accessible at http://5fur.genouest.org.

酿酒酵母是研究外界因素对细胞分裂和应激反应影响的极佳模型。几十年来,5-氟尿嘧啶(5-FU)一直被用于治疗实体瘤。这种药物最初是用来干扰 DNA 复制的,但后来发现它也能通过 RNA 依赖过程发挥抗增殖作用。由于 5-FU 可抑制酵母和哺乳动物体内 3'-5'- 外切核酸酶 Rrp6 的活性,因此早先的研究比较了二倍体同步酵母细胞中 5-FU 处理和 RRP6 缺失在转录组水平上的效果。为了便于解读表达数据,我们开发了一个改进的 5-FUR RNA 表达查看器。用户可以通过基因组坐标、mRNA 的系统或标准名称以及 Xrn1 依赖性、稳定、隐性和减数分裂未注释转录本(XUT、SUT、CUT 和 MUT)来获取信息。归一化对数 2 转换或线性数据可以填充图、线图或彩色热图的形式显示。这些表达数据对研究细胞周期调控、减数分裂基因的有丝分裂抑制、5-FU 处理和 Rrp6 缺乏对转录组的影响以及编码重叠转录本的有义/无义位点的表达谱等过程很有帮助。查看器可从 http://5fur.genouest.org 上获取。
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引用次数: 0
Exploring Saccharomycotina Yeast Ecology Through an Ecological Ontology Framework. 通过生态本体框架探索酵母生态学
IF 2.6 4区 生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-09-18 DOI: 10.1002/yea.3981
Marie-Claire Harrison,Dana A Opulente,John F Wolters,Xing-Xing Shen,Xiaofan Zhou,Marizeth Groenewald,Chris Todd Hittinger,Antonis Rokas,Abigail Leavitt LaBella
Yeasts in the subphylum Saccharomycotina are found across the globe in disparate ecosystems. A major aim of yeast research is to understand the diversity and evolution of ecological traits, such as carbon metabolic breadth, insect association, and cactophily. This includes studying aspects of ecological traits like genetic architecture or association with other phenotypic traits. Genomic resources in the Saccharomycotina have grown rapidly. Ecological data, however, are still limited for many species, especially those only known from species descriptions where usually only a limited number of strains are studied. Moreover, ecological information is recorded in natural language format limiting high throughput computational analysis. To address these limitations, we developed an ontological framework for the analysis of yeast ecology. A total of 1,088 yeast strains were added to the Ontology of Yeast Environments (OYE) and analyzed in a machine-learning framework to connect genotype to ecology. This framework is flexible and can be extended to additional isolates, species, or environmental sequencing data. Widespread adoption of OYE would greatly aid the study of macroecology in the Saccharomycotina subphylum.
酵母亚门中的酵母菌遍布全球不同的生态系统。酵母研究的一个主要目的是了解生态性状的多样性和进化,如碳代谢广度、与昆虫的关系和嗜仙人掌性。这包括研究生态性状的各个方面,如遗传结构或与其他表型性状的关联。酵母菌属的基因组资源增长迅速。然而,许多物种的生态学数据仍然有限,尤其是那些仅从物种描述中获知的物种,通常只研究有限数量的菌株。此外,以自然语言格式记录的生态信息也限制了高通量计算分析。为了解决这些限制,我们开发了一个用于分析酵母生态学的本体论框架。酵母环境本体(OYE)共添加了 1,088 株酵母菌株,并通过机器学习框架进行分析,将基因型与生态学联系起来。该框架非常灵活,可以扩展到更多的分离物、物种或环境测序数据。广泛采用 OYE 将大大有助于酵母亚门的宏观生态学研究。
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引用次数: 0
Improving an Alternative Glycerol Catabolism Pathway in Yarrowia lipolytica to Enhance Erythritol Production 改进脂肪分解酵母中的另一种甘油分解途径以提高赤藓糖醇产量
IF 2.6 4区 生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-09-12 DOI: 10.1002/yea.3980
Feng Liu, Jing‐Tao Tian, Ya‐Ting Wang, Lingxuan Zhao, Zhijie Liu, Jun Chen, Liu‐Jing Wei, Patrick Fickers, Qiang Hua
Engineering the glycerol‐3‐phosphate pathway could enhance erythritol production by accelerating glycerol uptake. However, little work has been conducted on the alternative dihydroxyacetone (DHA) pathway in Yarrowia lipolytica. Herein, this route was identified and characterized in Y. lipolytica by metabolomic and transcriptomic analysis. Moreover, the reaction catalyzed by dihydroxyacetone kinase encoded by dak2 was identified as the rate‐limiting step. By combining NHEJ‐mediated insertion mutagenesis with a push‐and‐pull strategy, Y. lipolytica strains with high‐yield erythritol synthesis from glycerol were obtained. Screening of a library of insertion mutants allows the identification of a mutant with fourfold increased erythritol production. Overexpression of DAK2 and glycerol dehydrogenase GCY3 together with gene encoding transketolase and transaldolase from the nonoxidative part of the pentose phosphate pathway led to a strain with further increased productivity with a titer of 53.1 g/L and a yield 0.56 g/g glycerol, which were 8.1‐ and 4.2‐fold of starting strain.
对甘油-3-磷酸途径进行工程改造可通过加速甘油吸收来提高赤藓糖醇的产量。然而,人们对脂肪分解亚罗威亚菌中的二羟基丙酮(DHA)替代途径研究甚少。本文通过代谢组学和转录组学分析,确定并描述了脂肪溶解酵母中的这一途径。此外,由 dak2 编码的二羟丙酮激酶催化的反应被确定为限速步骤。通过将 NHEJ 介导的插入突变与推拉策略相结合,获得了能从甘油中高产合成赤藓糖醇的溶脂芽孢杆菌菌株。通过筛选插入突变体文库,确定了赤藓糖醇产量提高四倍的突变体。过表达 DAK2 和甘油脱氢酶 GCY3 以及磷酸戊糖途径非氧化部分的反酮醇酶和反醛醇酶基因,可进一步提高菌株的产量,滴度为 53.1 克/升,甘油产量为 0.56 克/克,分别是起始菌株的 8.1 倍和 4.2 倍。
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引用次数: 0
Rapid Fluorescence Assay for Polyphosphate in Yeast Extracts Using JC‐D7 利用 JC-D7 对酵母提取物中的多聚磷酸盐进行快速荧光测定
IF 2.6 4区 生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-09-12 DOI: 10.1002/yea.3979
Alexander Deitert, Jana Fees, Anna Mertens, Duc Nguyen Van, Maria Maares, Hajo Haase, Lars Mathias Blank, Claudia Keil
Polyphosphate (polyP) is an intriguing molecule that is found in almost any organism, covering a multitude of cellular functions. In industry, polyP is used due to its unique physiochemical properties, including pH buffering, water binding, and bacteriostatic activities. Despite the importance of polyP, its analytics is still challenging, with the gold standard being 31P NMR. Here, we present a simple staining method using the fluorescent dye JC‐D7 for the semi‐quantitative polyP evaluation in yeast extracts. Notably, fluorescence response was affected by polyP concentration and polymer chain length in the 0.5–500 µg/mL polyP concentration range. Hence, for polyP samples of unknown chain compositions, JC‐D7 cannot be used for absolute quantification. Fluorescence of JC‐D7 was unaffected by inorganic phosphate up to 50 mM. Trace elements (FeSO4 > CuSO4 > CoCl2 > ZnSO4) and toxic mineral salts (PbNO3 and HgCl2) diminished polyP–induced JC‐D7 fluorescence, affecting its applicability to samples containing polyP–metal complexes. The fluorescence was only marginally affected by other parameters, such as pH and temperature. After validation, this simple assay was used to elucidate the degree of polyP production by yeast strains carrying gene deletions in (poly)phosphate homeostasis. The results suggest that staining with JC‐D7 provides a robust and sensitive method for detecting polyP in yeast extracts and likely in extracts of other microbes. The simplicity of the assay enables high‐throughput screening of microbes to fully elucidate and potentially enhance biotechnological polyP production, ultimately contributing to a sustainable phosphorus utilization.
聚磷酸盐(polyP)是一种有趣的分子,几乎存在于所有生物体内,具有多种细胞功能。在工业领域,聚磷酸酯因其独特的理化特性(包括 pH 缓冲、水结合和抑菌作用)而被广泛使用。尽管 polyP 十分重要,但对其进行分析仍然具有挑战性,目前的黄金标准是 31P NMR。在此,我们介绍一种使用荧光染料 JC-D7 的简单染色方法,用于对酵母提取物中的 polyP 进行半定量评估。值得注意的是,在 0.5-500 µg/mL 的 polyP 浓度范围内,荧光反应受 polyP 浓度和聚合物链长度的影响。因此,对于未知链组成的 polyP 样品,JC-D7 不能用于绝对定量。JC-D7 的荧光不受高达 50 mM 的无机磷酸盐的影响。微量元素(FeSO4 > CuSO4 > CoCl2 > ZnSO4)和有毒矿物盐(PbNO3 和 HgCl2)会减弱多聚物诱导的 JC-D7 荧光,从而影响其对含有多聚物-金属复合物的样品的适用性。荧光受 pH 值和温度等其他参数的影响很小。经过验证后,这种简单的检测方法被用于阐明携带(多)磷酸盐平衡基因缺失的酵母菌株产生多聚磷酸盐的程度。结果表明,JC-D7 染色法为检测酵母提取物中的 polyP 以及其他微生物提取物中的 polyP 提供了一种可靠而灵敏的方法。该检测方法简便易行,可对微生物进行高通量筛选,以全面阐明并提高生物技术生产多聚磷酸盐的潜力,最终促进磷的可持续利用。
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引用次数: 0
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