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Novel multifunctional plant growth-promoting bacteria isolated from the oil palm rhizosphere under long-term organic matter application 在长期施用有机物的条件下,从油棕根瘤菌中分离出的新型多功能植物生长促进菌。
IF 2.3 4区 生物学 Q3 BIOTECHNOLOGY & APPLIED MICROBIOLOGY Pub Date : 2024-08-06 DOI: 10.1016/j.jbiosc.2024.07.008
Fandi Hidayat , Rizki Desika Putri Pane , Fadilla Sapalina , Eka Listia , Winarna , Muhammad Edwin Syahputra Lubis , Mugihito Oshiro , Kenji Sakai , Yukihiro Tashiro
Most agricultural products are presently cultivated on marginal lands with poor soil properties and unfavorable environmental conditions (diseases and abiotic stresses), which can threaten plant growth and yield. Plant growth-promoting bacteria (PGPB) are beneficial bacteria that promote plant growth and biomass and act as biocontrols against diseases and stress. However, most isolated PGPBs have a single function and low survival rates owing to their limited growth behaviors. In this study, we isolated multifunctional PGPB from oil palm rhizosphere, quantitatively measured their activities, and evaluated their effectiveness in Brassica rapa (Komatsuna) cultivation. This is the first study to report the isolation of three multifunctional PGPB strains with ammonium production, phosphate-potassium-silicate solubilization, and indole-3-acetic acid (IAA) production from the oil palm rhizosphere, namely Kosakonia oryzendophytica AJLB38, Enterobacter quasimori AJTS77, and Lelliottia jeotgali AJTS83. Additionally, these strains showed antifungal activity against the oil palm pathogen Ganoderma boninense. These strains grow under high temperature, acidic and alkaline pH, and high salt concentration, which would result in their proliferation in various environmental conditions. The cultivation experiments revealed these strains improved the growth and biomass with half the dosage of chemical fertilizer application, which was not significantly different to the full dosage. Furthermore, the overall plant growth-promoting activities in quantitative assays and overall B. rapa growth in cultivation experiments were statistically correlated, which could contribute to the prediction of plant growth promotion without plant cultivation experiments. Thus, the selected PGPB could be valuable as a biofertilizer to improve soil health and quality and promote agricultural sustainability.
目前,大多数农产品都是在土壤贫瘠、环境条件不利(病害和非生物胁迫)的贫瘠土地上种植的,这可能会威胁到植物的生长和产量。植物生长促进菌(PGPB)是促进植物生长和生物量的有益细菌,也是抵御病害和胁迫的生物控制菌。然而,由于其生长行为有限,大多数分离出的 PGPB 功能单一且存活率低。在这项研究中,我们从油棕根瘤菌中分离出了多功能 PGPB,定量测定了它们的活性,并评估了它们在甘蓝型油菜(小松菜)栽培中的有效性。这是首次报道从油棕根瘤菌中分离出三种多功能 PGPB 菌株,即 Kosakonia oryzendophytica AJLB38、Enterobacter quasimori AJTS77 和 Lelliottia jeotgali AJTS83,它们具有产铵、磷酸-钾-硅酸盐增溶和产生吲哚-3-乙酸(IAA)的功能。此外,这些菌株对油棕病原体 Ganoderma boninense 具有抗真菌活性。这些菌株可在高温、酸碱 pH 值和高盐浓度条件下生长,因此可在各种环境条件下增殖。栽培实验表明,这些菌株在化肥施用量减半的情况下就能改善生长和生物量,与施用全量化肥相比没有显著差异。此外,定量检测中的总体植物生长促进活性与栽培实验中的蚕豆总体生长情况在统计学上存在相关性,这有助于在不进行植物栽培实验的情况下预测植物的生长促进情况。因此,所选的 PGPB 可作为生物肥料改善土壤健康和质量,促进农业可持续发展。
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引用次数: 0
Individual evaluation of nitrite and free nitrous acid inhibition on anammox activity 亚硝酸盐和游离亚硝酸对 Anammox 活性抑制作用的单独评估。
IF 2.3 4区 生物学 Q3 BIOTECHNOLOGY & APPLIED MICROBIOLOGY Pub Date : 2024-08-05 DOI: 10.1016/j.jbiosc.2024.06.010
Koya Hirose , Takashi Kondo , Yayoi Saito , Kazuichi Isaka

The anammox reaction simultaneously utilizes ammonia and nitrite as substrates; however, high nitrite concentrations act as strong inhibitors of the reaction. In this study, inhibition by NO2 and free nitrous acid (FNA) was separately evaluated in continuous feeding tests using different biomass carriers. The influent NO2 concentration was increased under pH 7.6, where FNA is less likely to affect anammox activity. A continuous test using polyethylene glycol (PEG) gel carriers containing immobilized anammox bacteria showed that the inhibition ratio was 13% when the NO2-N concentration in the reactor was 350 mg L−1 (FNA ≤0.06 mg L−1). The relationship between NO2 concentration in the reactor and inhibition ratio increased linearly. Evaluation of the inhibitory effect of FNA by increasing the influent NO2 concentration at pH 6.4, where FNA is easily formed, demonstrated that the relationship between FNA and inhibition ratio could be fitted to a sigmoid curve, and the 50% inhibitory concentration (IC50) of FNA was 0.88 mg L−1. A similar test performed using polyvinyl alcohol carriers containing anammox bacteria on their surface showed the same trend as the PEG gel carriers, with the IC50 for FNA at 0.70 mg L−1. These results indicate that the inhibitory effect of FNA on anammox activity was greater than that of NO2. The evaluation of these two factors helped identify important operational indicators of the stable application of anammox processes.

氨氧化反应同时利用氨和亚硝酸盐作为底物;然而,高浓度的亚硝酸盐会对该反应产生强烈的抑制作用。本研究使用不同的生物质载体,在连续进料试验中分别评估了 NO2- 和游离亚硝酸(FNA)的抑制作用。在 pH 值为 7.6 的条件下,进水的 NO2- 浓度增加,此时 FNA 对 anammox 活性的影响较小。使用含有固定化厌氧菌的聚乙二醇(PEG)凝胶载体进行的连续试验表明,当反应器中的 NO2-N 浓度为 350 mg L-1 时,抑制率为 13%(FNA ≤0.06 mg L-1)。反应器中的 NO2-浓度与抑菌率之间呈线性关系。在容易形成 FNA 的 pH 6.4 条件下,通过增加进水 NO2- 浓度来评估 FNA 的抑制作用,结果表明 FNA 与抑制比之间的关系可以拟合成一条 sigmoid 曲线,FNA 的 50% 抑制浓度 (IC50) 为 0.88 mg L-1。使用表面含有厌氧菌的聚乙烯醇载体进行的类似测试显示出与 PEG 凝胶载体相同的趋势,FNA 的 IC50 为 0.70 mg L-1。这些结果表明,FNA 对 Anammox 活性的抑制作用大于 NO2-。对这两个因素的评估有助于确定氨氧化工艺稳定应用的重要操作指标。
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引用次数: 0
Physiological responses contributing to multiple stress tolerance in Pichia kudriavzevii with potential enhancement for ethanol fermentation 有助于提高 Pichia kudriavzevii 对多种胁迫耐受性的生理反应,并有可能提高其乙醇发酵能力。
IF 2.3 4区 生物学 Q3 BIOTECHNOLOGY & APPLIED MICROBIOLOGY Pub Date : 2024-08-03 DOI: 10.1016/j.jbiosc.2024.07.012
Pongsanat Pongcharoen , Wittaya Tawong , Wanwarang Pathaichindachote , Weerawan Rod–in

Economically feasible ethanol production requires efficient hydrolysis of lignocellulosic biomass and high–temperature processing to enable simultaneous saccharification and fermentation. During the lignocellulolysic hydrolysate, the yeast must encounter with a multiple of inhibitors such as heat and furfural. To solve this problem, a potential fermentative yeast strain that tolerated simultaneous multistress and enhance ethanol concentration was investigated. Twenty yeast isolates were classified into two major yeast species, namely Pichia kudriavzevii (twelve isolates) and Candida tropicalis (eight isolates). All P. kudriavzevii isolates were able to grow at high temperature (45 °C) and exhibited stress tolerance toward furfural. Among P. kudriavzevii isolates, NUCG–S3 presented the highest specific growth rate under each stress condition of heat and furfural, and multistress. Morphological changes in P. kudriavzevii isolates (NUCG–S2, NUCG–S3, NUKL–P1, NUKL–P3, and NUOR–J1) showed alteration in mean cell length and width compared to the non–stress condition. Ethanol production by glucose was also determined. The yeast strain, NUCG–S3, gave the highest ethanol concentrations at 99.46 ± 0.82, 62.23 ± 0.96, and 65.80 ± 0.62 g/l (P < 0.05) under temperature of 30 °C, 40 °C, and 42 °C, respectively. The tolerant isolated yeast NUCG–S3 achieved ethanol production of 53.58 ± 3.36 and 48.06 ± 3.31 g/l (P < 0.05) in the presence of 15 mM furfural and multistress (42 °C with 15 mM furfural), respectively. Based on the results of the present study, the novel thermos and furfural-tolerant yeast strain P. kudriavzevii NUCG–S3 showed promise as a highly proficient yeast for high–temperature ethanol fermentation.

经济可行的乙醇生产需要对木质纤维素生物质进行高效水解和高温处理,以便同时进行糖化和发酵。在木质纤维素水解过程中,酵母必须遇到高温和糠醛等多种抑制剂。为解决这一问题,研究人员研究了一种能同时承受多重压力并提高乙醇浓度的潜在发酵酵母菌株。20 株酵母分离物被分为两大酵母种类,即 Pichia kudriavzevii(12 株)和 Candida tropicalis(8 株)。所有 P. kudriavzevii 分离物都能在高温(45 °C)下生长,并表现出对糠醛的应激耐受性。在 P. kudriavzevii 分离物中,NUCG-S3 在高温、糠醛和多重胁迫条件下的特定生长率最高。与非胁迫条件相比,P. kudriavzevii 分离物(NUCG-S2、NUCG-S3、NUKL-P1、NUKL-P3 和 NUOR-J1)的形态变化表现为平均细胞长度和宽度的改变。还测定了葡萄糖产生乙醇的情况。酵母菌株 NUCG-S3 在温度为 30 ℃、40 ℃ 和 42 ℃ 的条件下,乙醇浓度最高,分别为 99.46 ± 0.82、62.23 ± 0.96 和 65.80 ± 0.62 克/升(P < 0.05)。耐受性分离酵母 NUCG-S3 在 15 mM 糠醛和多重胁迫(42 °C、15 mM 糠醛)条件下的乙醇产量分别为 53.58 ± 3.36 和 48.06 ± 3.31 g/l(P < 0.05)。根据本研究的结果,新型耐高温、耐糠醛酵母菌株 P. kudriavzevii NUCG-S3 被认为是一种非常适合高温乙醇发酵的酵母菌。
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引用次数: 0
Regioselective photocyclodimerization of 2-anthracenecarboxylic acid through ATP hydrolysis-driven conformational change using simulation prediction-designed GroEL mutant 利用模拟预测设计的 GroEL 突变体,通过 ATP 水解驱动构象变化,实现 2-蒽羧酸的区域选择性光环二聚化。
IF 2.3 4区 生物学 Q3 BIOTECHNOLOGY & APPLIED MICROBIOLOGY Pub Date : 2024-08-02 DOI: 10.1016/j.jbiosc.2024.07.002
Masaki Nishijima , Kota Kobayashi , Megumi Masuda-Endo , Hiromi Yoda , Ayumi Koike-Takeshita

GroEL, a chaperone protein responsible for peptide and denatured protein folding, undergoes substantial conformational changes driven by ATP binding and hydrolysis during folding. Utilizing these conformational changes, we demonstrated the GroEL-mediated regioselective photocyclodimerization of 2-anthracenecarboxylic acid (AC) using ATP hydrolysis as an external stimulus. We designed and prepared an optimal GroEL mutant to employ in a docking simulation that has been actively used in recent years. Based on the large difference in the motif of hydrogen bonds between AC and GroEL mutant compared with the wild-type, we predicted that GroELMEL, in which the 307‒309th amino acid residues were mutated to Ala, could alter the orientation of bound AC in GroEL. The GroELMEL-mediated photocyclodimerization of AC can be used for regioselective inversion upon ATP addition to a moderate extent.

GroEL是一种负责多肽和变性蛋白质折叠的伴侣蛋白,在折叠过程中会因ATP的结合和水解而发生巨大的构象变化。利用这些构象变化,我们证明了以 ATP 水解为外部刺激,GroEL 介导了 2-蒽羧酸(AC)的区域选择性光环二聚化。我们设计并制备了一种最佳的 GroEL 突变体,并将其用于近年来一直在积极使用的对接模拟中。与野生型相比,AC 与 GroEL 突变体之间的氢键图案存在很大差异,基于此,我们预测将第 307-309 个氨基酸残基突变为 Ala 的 GroELMEL 可改变 GroEL 中结合的 AC 的取向。GroELMEL 介导的 AC 光环二聚化可在添加 ATP 时适度用于区域选择性反转。
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引用次数: 0
Peptide array screening with anti-GLP-1 monoclonal antibody: Discovery of cysteine-containing DPP-IV inhibitory peptides 用抗 GLP-1 单克隆抗体进行肽阵列筛选:发现含半胱氨酸的 DPP-IV 抑制肽。
IF 2.3 4区 生物学 Q3 BIOTECHNOLOGY & APPLIED MICROBIOLOGY Pub Date : 2024-07-31 DOI: 10.1016/j.jbiosc.2024.07.001
Masaki Kurimoto , Naoki Yuda , Masayoshi Tanaka , Miyuki Tanaka , Mina Okochi

Inhibition of dipeptidyl peptidase IV (DPP-IV) is an effective pharmacotherapy for the management of type 2 diabetes. Recent findings have suggested that various dietary proteins can serve as precursors to peptides that inhibit DPP-IV. Although several DPP-IV inhibitory peptides derived from food materials have been reported, more effective inhibitory peptides remain to be discovered. This study aimed to identify potent DPP-IV inhibitory peptides that earlier approaches had overlooked by employing a screening method that combined peptide arrays and neutralizing antibodies. Octa-peptides covering the complete amino acid sequences of four casein proteins and two whey proteins were synthesized on arrays via a solid-phase method. These peptides were then reacted with a monoclonal antibody specifically engineered to recognize glucagon-like peptide 1 (GLP-1), a substrate of DPP-IV. The variable region of the anti-GLP-1 monoclonal antibody is utilized to mimic the substrate-binding region of DPP-IV, enabling the antibody to bind to peptides that interact with DPP-IV. Based on this feature, 26 peptides were selected as DPP-IV inhibitory peptide candidates, 11 of which showed strong DPP-IV inhibitory activity. Five of these peptides consistently contained cysteines positioned two to four residues from the N-terminus. Treatment with disulfide formation decreased the DPP-IV inhibitory activity of these cysteine-containing peptides, while the inhibitory activity of α-lactalbumin hydrolysates increased with reducing treatment. These results revealed that the thiol group is important for DPP-IV inhibitory activity. This study provides a useful screen for DPP-IV inhibitory peptides and indicates the importance of reductive cysteine residues within DPP-IV inhibitory peptides.

抑制二肽基肽酶 IV(DPP-IV)是治疗 2 型糖尿病的一种有效药物疗法。最近的研究结果表明,各种膳食蛋白质可以作为抑制 DPP-IV 的肽的前体。虽然已有一些从食物中提取的 DPP-IV 抑制肽的报道,但更有效的抑制肽仍有待发现。本研究采用了一种结合肽阵列和中和抗体的筛选方法,旨在发现早期方法忽略的强效 DPP-IV 抑制肽。研究人员通过固相方法在肽阵列上合成了涵盖四种酪蛋白和两种乳清蛋白完整氨基酸序列的八胜肽。然后将这些肽与专门设计用于识别胰高血糖素样肽 1(GLP-1)(DPP-IV 的底物)的单克隆抗体反应。抗 GLP-1 单克隆抗体的可变区被用来模拟 DPP-IV 的底物结合区,使抗体能与 DPP-IV 相互作用的肽结合。根据这一特点,26 种肽被选为候选的 DPP-IV 抑制肽,其中 11 种具有很强的 DPP-IV 抑制活性。这些肽中有 5 种始终含有半胱氨酸,位于 N 端 2 到 4 个残基的位置。二硫化物的形成处理降低了这些含半胱氨酸肽的 DPP-IV 抑制活性,而 α-乳白蛋白水解物的抑制活性则随着还原处理的进行而增加。这些结果表明,硫醇基对 DPP-IV 抑制活性非常重要。这项研究为筛选 DPP-IV 抑制肽提供了有用的方法,并表明了还原性半胱氨酸残基在 DPP-IV 抑制肽中的重要性。
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引用次数: 0
Isolation of arginine deiminase system-deficient mutants of Tetragenococcus halophilus using arginine analog canavanine 利用精氨酸类似物卡纳瓦宁分离嗜卤四源球菌精氨酸脱氨酶系统缺陷突变体
IF 2.3 4区 生物学 Q3 BIOTECHNOLOGY & APPLIED MICROBIOLOGY Pub Date : 2024-07-29 DOI: 10.1016/j.jbiosc.2024.07.006
Takura Wakinaka , Jun Watanabe , Yoshinobu Mogi

Arginine deimination by Tetragenococcus halophilus, a halophilic lactic acid bacterium, is an undesirable reaction in soy sauce brewing because it is responsible for the production of ethyl carbamate, a potential carcinogen. Therefore, arginine deiminase system-deficient mutants have been generated and used as starter cultures. However, the pre-existing screening method for arginine deiminase system-deficient mutants was time consuming. To reduce the burden of this screening process, we established a method to isolate mutants incapable of arginine deimination using the arginine analog canavanine. Strains lacking arginine deiminase system were less sensitive to canavanine than wild type strain, which is likely because arginine deiminase consumes arginine in the cytoplasm and increases the relative concentration of canavanine in the cells and enhances its toxicity. This report provides an industrially useful method to efficiently obtain arginine deiminase system-deficient mutants.

嗜卤乳酸菌(Tetragenococcus halophilus)的精氨酸脱氨是酱油酿造中的一个不良反应,因为它会产生氨基甲酸乙酯(一种潜在的致癌物质)。因此,精氨酸脱氨酶系统缺陷突变体被生成并用作起始培养物。然而,现有的精氨酸脱氨酶系统缺陷突变体筛选方法非常耗时。为了减轻这一筛选过程的负担,我们建立了一种方法,利用精氨酸类似物卡那瓦宁来分离不能进行精氨酸脱氨的突变体。与野生型菌株相比,缺乏精氨酸脱氨酶系统的菌株对卡纳瓦宁的敏感性较低,这可能是因为精氨酸脱氨酶消耗了细胞质中的精氨酸,增加了细胞中卡纳瓦宁的相对浓度,增强了其毒性。本报告提供了一种有效获得精氨酸脱氨酶系统缺陷突变体的工业实用方法。
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引用次数: 0
Rational design approach to improve the solubility of the β-sandwich domain 1 of a thermophilic protein 提高嗜热蛋白质β-三明治结构域1溶解度的合理设计方法。
IF 2.3 4区 生物学 Q3 BIOTECHNOLOGY & APPLIED MICROBIOLOGY Pub Date : 2024-07-29 DOI: 10.1016/j.jbiosc.2024.06.009
Chukwuebuka M. Ononugbo , Yusaku Shimura , Noriko Yamano-Adachi , Takeshi Omasa , Yuichi Koga

The β-sandwich domain 1 (SD1) of islandisin is a stable thermophilic protein with surface loops that can be redesigned for specific target binding, architecturally comparable to the variable domain of immunoglobulin (IgG). SD1's propensity to aggregate due to incorrect folding and subsequent accumulation in Escherichia coli inclusion bodies limits its use in biotechnological applications. We rationally designed SD1 for improved variants that were expressed in soluble forms in E. coli while maintaining the intrinsic thermal stability of the protein (melting temperature (Tm) = 73). We used FoldX's ΔΔG predictions to find beneficial mutations and aggregation-prone regions (APRs) using Tango. The S26K substitution within protein core residues did not affect protein stability. Among the soluble mutants studied, the S26K/Q91P combination significantly improved the expression and solubility of SD1. We also examined the effects of the surface residue, pH, and concentration on the solubility of SD1. We showed that the surface polarity of proteins had little or no effect on solubility, whereas surface charges played a substantial role. The storage stability of several SD1 variants was impaired at pH values near their isoelectric point, and pH levels resulting in highly charged groups. We observed that mutations that create an uneven distribution of charged groups on the SD1 surface could enhance protein solubility by eliminating favorable protein–protein surface charge interactions. Our findings suggest that SD1 is mutationally tolerant to new functionalities, thus providing a novel perspective for the application of rational design to improve the solubility of targeted proteins.

海岛素的β-三明治结构域1(SD1)是一种稳定的嗜热蛋白质,其表面环路可根据特定目标结合而重新设计,在结构上类似于免疫球蛋白(IgG)的可变结构域。由于折叠不正确,SD1 容易聚集,随后在大肠杆菌包涵体中积累,这限制了它在生物技术应用中的使用。我们合理地设计了 SD1 的改良变体,这些变体可在大肠杆菌中以可溶形式表达,同时保持蛋白质的内在热稳定性(熔点温度 (Tm) = 73)。我们利用 FoldX 的 ΔΔG 预测,使用 Tango 找到了有益突变和易聚集区 (APR)。蛋白质核心残基中的 S26K 取代不会影响蛋白质的稳定性。在研究的可溶性突变体中,S26K/Q91P组合显著改善了SD1的表达和可溶性。我们还研究了表面残基、pH 值和浓度对 SD1 溶解性的影响。结果表明,蛋白质的表面极性对溶解度几乎没有影响,而表面电荷则对溶解度有很大影响。在接近等电点的 pH 值和导致高电荷基团的 pH 值水平下,几种 SD1 变体的储存稳定性受到影响。我们观察到,在 SD1 表面造成带电基团分布不均的突变可通过消除有利的蛋白质-蛋白质表面电荷相互作用来提高蛋白质的溶解度。我们的研究结果表明,SD1 对新的功能具有突变耐受性,从而为应用合理设计来提高目标蛋白质的溶解度提供了一个新的视角。
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引用次数: 0
Beneficial base substitutions in Escherichia coli fucO gene for enhancement of glycolic acid production 在大肠杆菌 fucO 基因中进行有益的碱基替换,以提高乙醇酸的产量。
IF 2.3 4区 生物学 Q3 BIOTECHNOLOGY & APPLIED MICROBIOLOGY Pub Date : 2024-07-29 DOI: 10.1016/j.jbiosc.2024.06.007
Mayu Nemoto , Wataru Muranushi , Chen Shuting , Yusuke Saito , Daisuke Sugimori , Miwa Yamada

Microbial production of glycolic acid (GA) from ethylene glycol is extensively used in a variety of industries because ethylene glycol is not only an inexpensive raw material but also the main component of industrial wastes. In this study, we produced GA from ethylene glycol using Escherichia coli overexpressing the endogenous 1,2-propanediol oxidoreductase (fucO) and lactaldehyde dehydrogenase (aldA) genes. To increase GA productivity, we screened a random mutant library generated using an error-prone polymerase chain reaction of fucO and obtained FucO mutants MF2-9 and MF6-9 with enhanced GA production in Lysogeny Broth medium containing 800 mM ethylene glycol. MF2-9 contained three amino acid substitutions (D23E, E222K, and G363S) and two synonymous mutations (coding DNA [c.] 93G > A and c.1131T > C) in fucO. MF6-9 contained one amino acid substitution (L377H) in FucO. An amino acid substitution (L377H) and a single synonymous mutation (c.1131T > C) in fucO contributed to the enhancement in GA production. Notably, cell lysates from E. coli harboring a synonymous mutation (c.1131T > C) or amino acid substitution (L377H) in fucO showed that only AldA activity was 1.3-fold higher than that of the cell lysate from E. coli harboring the wild-type fucO. We confirmed that c.1131T > C and L377H mutations increased aldA expression in E. coli. Analysis of mRNA levels and simulation of mRNA stabilization indicated that base substitutions at positions c.1130T, which corresponds to L377H amino acid substitution, and c.1131T increased aldA expression due to partial destabilization of the mRNA. These findings will be useful for the large-scale microbial production of GA from industrial waste.

微生物利用乙二醇生产乙醇酸(GA)被广泛应用于各行各业,因为乙二醇不仅是一种廉价的原材料,也是工业废料的主要成分。在本研究中,我们利用过表达内源 1,2-丙二醇氧化还原酶(fucO)和乳醛脱氢酶(aldA)基因的大肠杆菌从乙二醇中生产 GA。为了提高 GA 的产量,我们筛选了利用 fucO 的易错聚合酶链反应生成的随机突变体文库,并获得了 FucO 突变体 MF2-9 和 MF6-9,它们在含有 800 mM 乙二醇的溶菌培养基中的 GA 产量得到了提高。MF2-9 含有三个氨基酸替换(D23E、E222K 和 G363S)和两个同义突变(编码 DNA [c.] 93G > A 和 c.1131T > C)。MF6-9 在 FucO 中含有一个氨基酸替换(L377H)。FucO 中的一个氨基酸取代(L377H)和一个同义突变(c.1131T > C)导致了 GA 产量的增加。值得注意的是,fucO 中携带同义突变(c.1131T > C)或氨基酸替换(L377H)的大肠杆菌细胞裂解液显示,只有 AldA 活性比携带野生型 fucO 的大肠杆菌细胞裂解液高 1.3 倍。我们证实,c.1131T > C 和 L377H 突变增加了大肠杆菌中 aldA 的表达。对 mRNA 水平的分析和 mRNA 稳定性的模拟表明,c.1130T(相当于 L377H 氨基酸置换)和 c.1131T 位置的碱基置换会增加 aldA 的表达,这是由于 mRNA 的部分不稳定所致。这些发现将有助于从工业废物中大规模微生物生产 GA。
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引用次数: 0
Isolation of aflatoxin biosynthetic inhibitor from Chondrostereum purpureum mushroom culture filtrate 从软玉菇培养滤液中分离出黄曲霉毒素生物合成抑制剂。
IF 2.3 4区 生物学 Q3 BIOTECHNOLOGY & APPLIED MICROBIOLOGY Pub Date : 2024-07-26 DOI: 10.1016/j.jbiosc.2024.07.003
Yuya Matsuno , Naoki Endo , Kotomi Ueno , Atsushi Ishihara

Aflatoxins (AFs) are highly toxic mycotoxins produced by the fungi, Aspergillus flavus and Aspergillus parasiticus. AFs pose severe health risks owing to their acute toxicity and carcinogenic properties. The control of AF contamination remains significantly challenging despite the extensive efforts toward controlling it. Here, we investigated the potential of mushroom extracts as a source of AF biosynthetic inhibitors. The A. parasiticus mutant strain, NFRI-95, that accumulates an AF biosynthesis intermediate, norsolorinic acid, was used in the bioassay to detect the inhibitory activity against AF biosynthesis. The screening of 195 mushroom extracts revealed that the culture filtrate extract of Chondrostereum purpureum exhibited strong inhibitory activity against AF biosynthesis. Next, large-scale culturing of C. purpureum was performed to isolate the compounds accounting for the inhibitory activity. The culture filtrate was extracted with ethyl acetate, after which the active compound was isolated by silica gel column chromatography and preparative high performance liquid chromatography (HPLC). The active compound was identified as cyclo(Val–Pro) by spectroscopic analyses. Further, four stereoisomers of cyclo(Val–Pro) were synthesized by the condensation of the N-Boc derivatives of d- and l-valine with the methyl esters of d- and l-proline. The naturally isolated compound was identified as cyclo(l-Val-l-Pro) by comparing its retention time with those of synthetic compounds by chiral HPLC analysis and CD spectra. The IC50 value of cyclo(L-Val-L-Pro) was 2.4 mM, whereas the LD, DL, and DD isomers exhibited weaker activities, with IC50 values of >5 mM.

黄曲霉毒素(AFs)是由真菌黄曲霉和寄生曲霉产生的剧毒霉菌毒素。黄曲霉毒素具有急性毒性和致癌性,严重危害人们的健康。尽管人们在控制 AF 污染方面做出了大量努力,但控制 AF 污染仍面临巨大挑战。在这里,我们研究了蘑菇提取物作为一种 AF 生物合成抑制剂来源的潜力。生物测定中使用了寄生蝇突变菌株 NFRI-95,该菌株可积累甲酸萘酯生物合成中间体--萘甲酸,以检测其对甲酸萘酯生物合成的抑制活性。195 种蘑菇提取物的筛选结果表明,紫软骨蘑的培养滤液提取物对 AF 的生物合成具有很强的抑制活性。接下来,对紫软骨蘑进行了大规模培养,以分离出具有抑制活性的化合物。用乙酸乙酯萃取培养滤液,然后通过硅胶柱层析和制备型高效液相色谱法(HPLC)分离出活性化合物。通过光谱分析,活性化合物被确定为环(缬氨脯)。此外,通过 d-和 l-缬氨酸的 N-Boc衍生物与 d-和 l-脯氨酸的甲酯缩合,合成了四种环(缬氨脯)立体异构体。通过手性高效液相色谱分析和 CD 光谱,比较其保留时间和合成化合物的保留时间,确定了自然分离的化合物为环(l-缬氨酰-l-脯氨酸)。环(L-缬氨酰-L-脯氨酸)的 IC50 值为 2.4 mM,而 LD、DL 和 DD 异构体的活性较弱,IC50 值大于 5 mM。
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引用次数: 0
Isolated hair bacteria reveal different isolation possibilities under various conditions 分离出的毛发细菌显示了在不同条件下的不同分离可能性。
IF 2.3 4区 生物学 Q3 BIOTECHNOLOGY & APPLIED MICROBIOLOGY Pub Date : 2024-07-20 DOI: 10.1016/j.jbiosc.2024.06.003
Azusa Yamada , Yuri Nishi , Mei Noguchi , Kota Watanabe , Mugihito Oshiro , Kenji Sakai , Yukihiro Tashiro

Microorganisms are assumed to inhabit various environments and organisms, including the human body. The presence of more than 700 bacterial species on scalp hair has been reported through rRNA gene amplicon analysis. However, the biological properties of bacteria on the scalp hair (hair bacteria) and their functions are poorly understood as few hair bacteria have been isolated from hair in previous studies. This study aimed to isolate hair bacteria using standard media under 24 different conditions (including medium components, component concentrations, gelling agents, and atmospheric environments). Furthermore, we evaluated the possibility of isolating strains under these isolation conditions and examined the carbon metabolic ability of several predominantly isolated strains. A total of 63 bacterial species belonging to 27 genera were isolated from hair under 24 isolation conditions. The predominant bacterial species isolated from human hair in this study showed different carbon metabolic capabilities than those of the reference strains. In addition, isolation possibility was newly proposed to systematically evaluate the number of isolation conditions that could cultivate a bacterial species. Based on isolation possibility, the isolates were categorized into groups with a high number of isolation conditions (e.g., ≥25%; such as Staphylococcus) and those with a low number (e.g., ≤25%; such as Brachybacterium). These findings indicate the existence of easily isolated microorganisms and difficultly isolated microorganism from human hair.

微生物被认为栖息在包括人体在内的各种环境和生物体中。据报道,通过 rRNA 基因扩增片段分析,头皮头发上存在 700 多种细菌。然而,人们对头皮毛发上细菌(毛发细菌)的生物特性及其功能知之甚少,因为以往的研究很少从毛发中分离出毛发细菌。本研究旨在使用标准培养基,在 24 种不同条件下(包括培养基成分、成分浓度、胶凝剂和大气环境)分离头发细菌。此外,我们还评估了在这些分离条件下分离菌株的可能性,并考察了几种主要分离菌株的碳代谢能力。在 24 种分离条件下,共从头发中分离出 27 个属的 63 种细菌。与参考菌株相比,本研究从人类头发中分离出的主要细菌种类表现出不同的碳代谢能力。此外,还新提出了分离可能性,以系统评估可培养细菌物种的分离条件数量。根据分离可能性,分离菌株被分为高分离条件组(如≥25%;如葡萄球菌)和低分离条件组(如≤25%;如分支杆菌)。这些发现表明,人类头发中存在易分离微生物和难分离微生物。
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引用次数: 0
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Journal of bioscience and bioengineering
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