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Transglutaminase mediates the hardening of fish egg envelope produced by duplication of factor XIIIA gene during the evolution of Teleostei. 转谷氨酰胺酶在鱼类进化过程中介导了因子 XIIIA 基因复制产生的鱼卵包膜硬化。
IF 2.1 4区 生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-09-16 DOI: 10.1093/jb/mvae062
Shigeki Yasumasu, Miyuki Horie, Mayuko Horie, Kodai Sakuma, Chihiro Sato, Hikari Sato, Taiki Nakajima, Tatsuki Nagasawa, Mari Kawaguchi, Ichiro Iuchi

During the fertilization of fish eggs, the hardening of the egg envelope is mediated by transglutaminase (hTGase). After fertilization, TGase undergoes processing. We isolated hTGase from extracts of unfertilized and water-activated rainbow trout eggs. Rainbow trout hTGase (Rt-hTGase) appeared as an 80 kDa protein, and its processed form was 55 kDa. Their N-terminal amino acid sequences were nearly identical, suggesting processing in the C-terminal region. The specific activities were not significantly different, indicating that C-terminal processing does not activate the enzyme itself. We cloned the cDNA by reverse transcription polymerase chain reaction (RT-PCR) using degenerate primers followed by RACE-PCR. The deduced amino acid sequence of the cDNA was similar to that of factor XIII subunit A (FXIIIA). Molecular phylogenetic and gene syntenic analyses clearly showed that hTGase was produced by duplication of FXIIIA during the evolution to Teleostei. The 55 kDa processed form of Rt-hTGase is predominantly composed of an enzyme domain predicted from the amino acid sequence of the cDNA. It is hypothesized that the C-terminal domain of Rt-hTGase binds to egg envelope proteins, and that processing allows the enzyme to move freely within the egg envelope, increasing substrate-enzyme interaction and thereby accelerating hardening.

在鱼卵受精过程中,卵包膜的硬化是由转谷氨酰胺酶(hTGase)介导的。受精后,转谷氨酰胺酶会进行加工。我们从未受精虹鳟鱼卵和水活化虹鳟鱼卵的提取物中分离出了 hTGase。虹鳟鱼 hTGase(Rt-hTGase)呈 80 kDa 蛋白,其加工形式为 55 kDa。它们的 N 端氨基酸序列几乎相同,表明在 C 端区域进行了加工。它们的特异性活性没有明显差异,表明 C 端加工并没有激活酶本身。我们使用变性引物通过反转录聚合酶链反应(RT-PCR)克隆了 cDNA,然后进行了 RACE-PCR。cDNA 的推导氨基酸序列与因子 XIII 亚基 A(FXIIIA)相似。分子系统发育和基因同源分析清楚地表明,hTGase 是由 FXIIIA 在向长尾目进化过程中复制产生的。Rt-hTGase 的 55 kDa 加工形式主要由根据 cDNA 氨基酸序列预测的酶域组成。据推测,Rt-hTGase 的 C 端结构域可与卵包膜蛋白质结合,加工后的酶可在卵包膜内自由移动,增加底物与酶的相互作用,从而加速硬化。
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引用次数: 0
Neutral selection and clonal expansion during the development of colon cancer metastasis. 结肠癌转移发展过程中的中性选择和克隆扩增。
IF 2.1 4区 生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-09-03 DOI: 10.1093/jb/mvae044
Xuelian Lei, Daisuke Yamamoto, Hirotaka Kitamura, Kenji Kita, Noriyuki Inaki, Kazuhiro Murakami, Mizuho Nakayama, Hiroko Oshima, Masanobu Oshima

Intratumour heterogeneity has been shown to play a role in the malignant progression of cancer. The clonal evolution in primary cancer has been well studied, however, that in metastatic tumorigenesis is not fully understood. In this study, we established human colon cancer-derived organoids and investigated clonal dynamics during liver metastasis development by tracking barcode-labelled subclones. Long-term subclone co-cultures showed clonal drift, with a single subclone becoming dominant in the cell population. Interestingly, the selected subclones were not always the same, suggesting that clonal selection was not based on cell intrinsic properties. Furthermore, liver tumours developed by co-transplantation of organoid subclones into the immunodeficient mouse spleen showed a progressive drastic reduction in clonal diversity, and only one or two subclones predominated in the majority of large metastatic tumours. Importantly, selections were not limited to particular subclones but appeared to be random. A trend towards a reduction in clonal diversity was also found in liver metastases of multiple colour-labelled organoids of mouse intestinal tumours. Based on these results, we propose a novel mechanism of metastasis development, i.e. a subclone population of the disseminated tumour cells in the liver is selected by neutral selection during colonization and constitutes large metastatic tumours.

肿瘤内的异质性已被证明在癌症的恶性发展中发挥作用。尽管对原发性癌症中的克隆演化已有深入研究,但对转移性肿瘤发生过程中的克隆演化还不完全了解。在这项研究中,我们建立了人结肠癌器官组织,并通过追踪条形码标记的亚克隆研究了肝转移发展过程中的克隆动态。长期的亚克隆共培养显示出克隆漂移,单个亚克隆在细胞群中占据主导地位。有趣的是,被选择的亚克隆并不总是相同的,这表明克隆选择并非基于细胞的内在特性。此外,将类器官亚克隆共同移植到免疫缺陷小鼠脾脏而形成的肝脏肿瘤显示,克隆多样性逐渐急剧下降,在大多数大的转移性肿瘤中,只有一个或两个亚克隆占主导地位。重要的是,选择并不局限于特定的亚克隆,而似乎是随机的。在小鼠肠道肿瘤的多个彩色标记器官组织的肝转移瘤中也发现了克隆多样性减少的趋势。基于这些结果,我们提出了一种新的转移发展机制,即肝脏中扩散的肿瘤细胞的亚克隆群体在定殖过程中被中性选择,并构成大型转移性肿瘤。
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引用次数: 0
Onnamide A suppresses the severe acute respiratory syndrome-coronavirus 2 infection without inhibiting 3-chymotrypsin-like cysteine protease. 翁内酰胺 A 可抑制严重急性呼吸系统综合征-冠状病毒 2 感染,而不会抑制 3-糜蛋白酶样半胱氨酸蛋白酶。
IF 2.1 4区 生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-09-03 DOI: 10.1093/jb/mvae037
Yasuhiro Hayashi, Nanami Higa, Tetsuro Yoshida, Trianda Ayuning Tyas, Kanami Mori-Yasumoto, Mina Yasumoto-Hirose, Hideki Tani, Junichi Tanaka, Takahiro Jomori

Given the continuous emergence of new variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the development of new inhibitors is necessary to enhance clinical efficacy and increase the options for combination therapy for the coronavirus disease 2019. Because marine organisms have been a resource for the discovery of numerous bioactive molecules, we constructed an extract library of marine invertebrates collected from the Okinawa Islands. In this study, the extracts were used to identify antiviral molecules against SARS-CoV-2. Using a cytopathic effect (CPE) assay in VeroE6/TMPRSS2 cells, an extract from the marine sponge Theonella swinhoei was found to reduce virus-induced CPE. Eventually, onnamide A was identified as an antiviral compound in the extract using column chromatography and NMR analysis. Onnamide A inhibited several SARS-CoV-2 variant-induced CPEs in VeroE6/TMPRSS2 cells as well as virus production in the supernatant of infected cells. Moreover, this compound blocked the entry of SARS-CoV-2 pseudo-virions. Taken together, these results demonstrate that onnamide A suppresses SARS-CoV-2 infection, which may be partially related to entry inhibition, and is expected to be a candidate lead compound for the development of anti-SARS-CoV-2 drugs.

鉴于严重急性呼吸系统综合征冠状病毒 2(SARS-CoV-2)新变种的不断出现,有必要开发新的抑制剂,以提高临床疗效,增加 2019 年冠状病毒疾病联合疗法的选择。由于海洋生物是发现大量生物活性分子的资源,我们构建了一个从冲绳群岛采集的海洋无脊椎动物提取物库。在这项研究中,我们利用这些提取物来鉴定针对 SARS-CoV-2 的抗病毒分子。通过在 VeroE6/TMPRSS2 细胞中进行细胞病理效应(CPE)试验,发现海洋海绵 Theonella swinhoei 的提取物可减少病毒诱导的 CPE。通过柱层析和核磁共振分析,最终确定翁酰胺 A 是提取物中的一种抗病毒化合物。Onnamide A 能抑制 VeroE6/TMPRSS2 细胞中几种 SARS-CoV-2 变体诱导的 CPE 以及感染细胞上清液中病毒的产生。此外,这种化合物还能阻止 SARS-CoV-2 伪病毒的进入。综上所述,这些结果表明昂纳米德 A 能抑制 SARS-CoV-2 感染,部分原因可能与抑制病毒进入有关,有望成为开发抗 SARS-CoV-2 药物的候选先导化合物。
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引用次数: 0
Amino acid residues responsible for the different pH dependency of cell-specific ferredoxins in the electron transfer reaction with ferredoxin-NADP+ reductase from maize leaves. 在与玉米叶片中的铁氧还蛋白-NADP+还原酶进行电子转移反应时,细胞特异性铁氧还蛋白对 pH 值的依赖性不同的氨基酸残基。
IF 2.1 4区 生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-09-03 DOI: 10.1093/jb/mvae043
Yoko Kimata-Ariga, Hikaru Tanaka, Shunsuke Kuwano

In the chloroplast stroma, dynamic pH changes occur from acidic to alkaline in response to fluctuating light conditions. We investigated the pH dependency of the electron transfer reaction of ferredoxin-NADP+ reductase (FNR) with ferredoxin (Fd) isoproteins, Fd1 and Fd2, which are localized in mesophyll cells and bundle sheath cells, respectively, in the leaves of C4 plant maize. The pH-dependent profile of the electron transfer activity with FNR was quite different between Fd1 and Fd2, which was mainly explained by the opposite pH dependency of the Km value of these Fds for FNR. Replacement of the amino acid residue at position of 65 (D65N) and 78 (H78A) between the two Fds conferred different effect on their pH dependency of the Km value. Double mutations of the two residues between Fd1 and Fd2 (Fd1D65N/H78A and Fd2N65D/A78H) led to the mutual exchange of the pH dependency of the electron transfer activity. This exchange was mainly explained by the changes in the pH-dependent profile of the Km values. Therefore, the differences in Asp/Asn at position 65 and His/Ala at position 78 between Fd1 and Fd2 were shown to be the major determinants for their different pH dependency in the electron transfer reaction with FNR.

在叶绿体基质中,pH值会随着光照条件的变化而发生从酸性到碱性的动态变化。我们研究了铁氧还蛋白-NADP+还原酶(FNR)与铁氧还蛋白(Fd)异构体Fd1和Fd2的电子传递反应的pH依赖性。Fd1 和 Fd2 与 FNR 的电子传递活性随 pH 值变化的曲线截然不同,这主要是因为这两种 Fds 对 FNR 的 Km 值与 pH 值的依赖性相反。两个 Fds 之间 65(D65N)和 78(H78A)位氨基酸残基的置换对其 Km 值的 pH 依赖性产生了不同的影响。Fd1 和 Fd2 之间两个残基的双突变(Fd1D65N/H78A 和 Fd2N65D/A78H)导致电子转移活性的 pH 依赖性相互交换。这种交换的主要原因是 Km 值的 pH 依赖性曲线发生了变化。因此,Fd1 和 Fd2 在与 FNR 进行电子转移反应时,其 65 位的 Asp/Asn 和 78 位的 His/Ala 的差异被证明是导致其 pH 依赖性不同的主要决定因素。
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引用次数: 0
Comprehensive analysis of non-selective and selective autophagy in yeast atg mutants and characterization of autophagic activity in the absence of the Atg8 conjugation system. 全面分析酵母 atg 突变体的非选择性和选择性自噬,以及 Atg8 连接系统缺失时自噬活性的特征。
IF 2.1 4区 生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-09-03 DOI: 10.1093/jb/mvae042
Tamara Ginevskaia, Aleksei Innokentev, Kentaro Furukawa, Tomoyuki Fukuda, Manabu Hayatsu, Shun-Ichi Yamashita, Keiichi Inoue, Shinsuke Shibata, Tomotake Kanki

Most autophagy-related genes, or ATG genes, have been identified through studies using budding yeast. Although the functions of the ATG genes are well understood, the contributions of individual genes to non-selective and various types of selective autophagy remain to be fully elucidated. In this study, we quantified the activity of non-selective autophagy, the cytoplasm-to-vacuole targeting (Cvt) pathway, mitophagy, endoplasmic reticulum (ER)-phagy and pexophagy in all Saccharomyces cerevisiae atg mutants. Among the mutants of the core autophagy genes considered essential for autophagy, the atg13 mutant and mutants of the genes involved in the two ubiquitin-like conjugation systems retained residual autophagic functionality. In particular, mutants of the Atg8 ubiquitin-like conjugation system (the Atg8 system) exhibited substantial levels of non-selective autophagy, the Cvt pathway and pexophagy, although mitophagy and ER-phagy were undetectable. Atg8-system mutants also displayed intravacuolar vesicles resembling autophagic bodies, albeit at significantly reduced size and frequency. Thus, our data suggest that membranous sequestration and vacuolar delivery of autophagic cargo can occur in the absence of the Atg8 system. Alongside these findings, the comprehensive analysis conducted here provides valuable datasets for future autophagy research.

大多数自噬相关基因或 ATG 基因都是在利用芽殖酵母进行的研究中发现的。虽然 ATG 基因的功能已广为人知,但各个基因对非选择性自噬和各种选择性自噬的贡献仍有待全面阐明。在这项研究中,我们量化了所有酿酒酵母 atg 突变体中的非选择性自噬、细胞质到液泡靶向(Cvt)途径、有丝分裂、内质网(ER)自噬和pexophagy 的活性。在被认为对自噬至关重要的核心自噬基因突变体中,atg13 突变体和参与两个泛素样连接系统的基因突变体保留了残余的自噬功能。特别是 Atg8 泛素样连接系统(Atg8 系统)的突变体表现出大量的非选择性自噬、Cvt 途径和pexophagy,但丝裂噬和ER-噬却检测不到。Atg8 系统突变体也显示出类似自噬体的泡内囊泡,尽管其大小和频率明显降低。因此,我们的数据表明,在没有 Atg8 系统的情况下,自噬货物的膜螯合和液泡输送也会发生。除了这些发现,本文进行的综合分析还为未来的自噬研究提供了宝贵的数据集。
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引用次数: 0
Chondroitin sulfate liposome: clustering toward high functional efficiency. 硫酸软骨素脂质体:向高功能效率集聚。
IF 2.1 4区 生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-09-03 DOI: 10.1093/jb/mvae041
Tatsumasa Shioiri, Jun Tsuchimoto, Kaori Fukushige, Takao Takeuchi, Munekazu Naito, Hideto Watanabe, Nobuo Sugiura

Chondroitin sulfate (CS) is a linear polysaccharide chain of alternating residues of glucuronic acid (GlcA) and N-acetylgalactosamine (GalNAc), modified with sulfate groups. Based on the structure, CS chains bind to bioactive molecules specifically and regulate their functions. For example, CS whose GalNAc is sulfated at the C4 position, termed CSA, and CS whose GalNAc is sulfated at both C4 and C6 positions, termed CSE, bind to a malaria protein VAR2CSA and receptor type of protein tyrosine phosphatase sigma (RPTPσ), respectively, in a specific manner. Here, we modified CSA and CSE chains with phosphatidylethanolamine (PE) at a reducing end, attached them to liposomes containing phospholipids and generated CSA and CSE liposomes. The CS-PE was incorporated into the liposome particles efficiently. Inhibition ELISA revealed specific interaction of CSA and CSE with recombinant VAR2CSA and RPTPσ, respectively, more efficiently than CS chains alone. Furthermore, CSE liposome was specifically incorporated into RPTPσ-expressing HEK293T cells. These results indicate CS liposome as a novel and efficient drug delivery system, especially for CS-binding molecules.

硫酸软骨素(CS)是由葡萄糖醛酸(GlcA)和 N-乙酰半乳糖胺(GalNAc)交替残基组成的线性多糖链,并用硫酸基团修饰。根据其结构,CS 链可与生物活性分子特异性结合并调节其功能。例如,GalNAc 在 C4 位硫酸化的 CS 被称为 CSA,GalNAc 在 C4 和 C6 位硫酸化的 CS 被称为 CSE,它们分别以特定的方式与疟疾蛋白 VAR2CSA 和受体型蛋白酪氨酸磷酸酶 sigma(RPTPσ)结合。在这里,我们在还原端用磷脂酰乙醇胺(PE)修饰了 CSA 和 CSE 链,将它们连接到含有磷脂的脂质体上,生成了 CSA 和 CSE 脂质体。CS-PE 被有效地结合到脂质体颗粒中。抑制酶联免疫吸附试验表明,CSA 和 CSE 分别与重组 VAR2CSA 和 RPTPσ 的特异性相互作用比单独的 CS 链更有效。此外,CSE-脂质体还能特异性地结合到表达 RPTPσ 的 HEK293T 细胞中。这些结果表明 CSE 脂质体是一种新型高效的药物输送系统,尤其适用于 CS 结合分子。
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引用次数: 0
Selection and characterization of aptamers targeting the Vif-CBFβ-ELOB-ELOC-CUL5 complex. 以 Vif-CBFβ-ELOB-ELOC-CUL5 复合物为靶标的适配体的筛选和表征。
IF 2.1 4区 生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-09-03 DOI: 10.1093/jb/mvae040
Kazuyuki Kumagai, Keisuke Kamba, Takuya Suzuki, Yuto Sekikawa, Chisato Yuki, Michiaki Hamada, Kayoko Nagata, Akifumi Takaori-Kondo, Li Wan, Masato Katahira, Takashi Nagata, Taiichi Sakamoto

The viral infectivity factor (Vif) of human immunodeficiency virus 1 forms a complex with host proteins, designated as Vif-CBFβ-ELOB-ELOC-CUL5 (VβBCC), initiating the ubiquitination and subsequent proteasomal degradation of the human antiviral protein APOBEC3G (A3G), thereby negating its antiviral function. Whilst recent cryo-electron microscopy (cryo-EM) studies have implicated RNA molecules in the Vif-A3G interaction that leads to A3G ubiquitination, our findings indicated that the VβBCC complex can also directly impede A3G-mediated DNA deamination, bypassing the proteasomal degradation pathway. Employing the Systematic Evolution of Ligands by EXponential enrichment (SELEX) method, we have identified RNA aptamers with high affinity for the VβBCC complex. These aptamers not only bind to the VβBCC complex but also reinstate A3G's DNA deamination activity by inhibiting the complex's function. Moreover, we delineated the sequences and secondary structures of these aptamers, providing insights into the mechanistic aspects of A3G inhibition by the VβBCC complex. Analysis using selected aptamers will enhance our understanding of the inhibition of A3G by the VβBCC complex, offering potential avenues for therapeutic intervention.

人类免疫缺陷病毒 1 的病毒感染因子(Vif)会与宿主蛋白形成复合物,即 Vif-CBFβ-ELOB-ELOC-CUL5(VβBCC),启动人类抗病毒蛋白 APOBEC3G(A3G)的泛素化和随后的蛋白酶体降解,从而使其丧失抗病毒功能。最近的低温电子显微镜(cryo-EM)研究表明,RNA分子与Vif-A3G相互作用,导致A3G泛素化,而我们的研究结果表明,VβBCC复合物还能绕过蛋白酶体降解途径,直接阻碍A3G介导的DNA脱氨。利用配体的系统进化富集(SELEX)方法,我们发现了与 VβBCC 复合物具有高亲和力的 RNA 合体。这些适配体不仅能与 VβBCC 复合物结合,还能通过抑制该复合物的功能来恢复 A3G 的 DNA 脱氨活性。此外,我们还描绘了这些适配体的序列和二级结构,为深入了解 VβBCC 复合物抑制 A3G 的机理提供了线索。利用选定的适配体进行分析将加深我们对 VβBCC 复合物抑制 A3G 的理解,为治疗干预提供潜在的途径。
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引用次数: 0
Neurodegenerative diseases associated with the disruption of proteostasis and their therapeutic strategies using chemical chaperones. 与蛋白稳态破坏有关的神经退行性疾病及其利用化学伴侣的治疗策略。
IF 2.1 4区 生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-09-03 DOI: 10.1093/jb/mvae048
Takashi Sugiyama, Hideki Nishitoh

Aberrant proteostasis is thought to be involved in the pathogenesis of neurodegenerative diseases. Some proteostasis abnormalities are ameliorated by chaperones. Chaperones are divided into three groups: molecular, pharmacological and chemical. Chemical chaperones intended to alleviate stress in organelles, such as the endoplasmic reticulum (ER), are now being administered clinically. Of the chemical chaperones, 4-phenylbutyrate (4-PBA) has been used as a research reagent, and its mechanism of action includes chaperone effects and the inhibition of histone deacetylase. Moreover, it also binds to the B-site of SEC24 and regulates COPII-mediated transport from the ER. Although its therapeutic effect may not be strong, elucidating the mechanism of action of 4-PBA may contribute to the identification of novel therapeutic targets for neurodegenerative diseases.

蛋白稳态异常被认为与神经退行性疾病的发病机制有关。一些蛋白稳态异常可通过伴侣素得到改善。伴侣分为三类:分子伴侣、药理学伴侣和化学伴侣。化学伴侣旨在减轻细胞器(如内质网)的压力,目前已在临床上使用。在化学伴侣剂中,4-苯基丁酸酯(4-PBA)已被用作研究试剂,其作用机制包括伴侣效应和抑制组蛋白去乙酰化酶。此外,它还能与 SEC24 的 B 位点结合,调节 COPII 介导的从 ER 的转运。虽然 4-PBA 的治疗效果可能不强,但阐明其作用机制可能有助于确定神经退行性疾病的新治疗靶点。
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引用次数: 0
CDP-DAG synthesis by peripheral membrane-bound Tam41-type enzymes. 外周膜结合的 Tam41 型酶合成 CDP-DAG。
IF 2.1 4区 生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-09-03 DOI: 10.1093/jb/mvae046
Koji Okamoto

Cytidine diphosphate diacylglycerol (CDP-DAG) is a critical intermediate that is converted to multiple phospholipids in prokaryotes and eukaryotes. In budding yeast, CDP-DAG synthesis from cytidine triphosphate (CTP) and phosphatidic acid (PA) is catalyzed by the membrane-integrated protein Cds1 in the endoplasmic reticulum and the peripheral membrane-bound protein Tam41 in mitochondria. Although a recent study revealed that the fission yeast SpTam41 consists of a nucleotidyltransferase domain and a winged helix domain, forming an active-site pocket for CTP binding between the two domains together with a C-terminal amphipathic helix for membrane association, how CTP and Mg 2+, a most-favoured divalent cation, are accommodated with PA remains obscure. A more recent report by Kimura et al. (J. Biochem. 2022; 171:429-441) solved the crystal structure of FbTam41, a functional ortholog from a Firmicutes bacterium, with CTP-Mg 2+, successfully providing a detailed molecular view of CDP-DAG synthesis. In this commentary, our current understanding of Tam41-mediated reaction is discussed.

胞苷二磷酸二酰甘油(CDP-DAG)是原核生物和真核生物转化为多种磷脂的关键中间体。在芽殖酵母中,CDP-DAG 由三磷酸胞苷(CTP)和磷脂酸(PA)合成,由内质网中的膜整合蛋白 Cds1 和线粒体中的外周膜结合蛋白 Tam41 催化。尽管最近的一项研究发现,裂殖酵母的 SpTam41 由一个核苷酸转移酶结构域和一个翼状螺旋结构域组成,在两个结构域之间形成一个用于结合 CTP 的活性位点口袋,C-末端的两性螺旋用于与膜结合,但 CTP 和 Mg2+(最喜欢的二价阳离子)如何与 PA 相容仍不清楚。最近,Kimura 等人的报告(J. Biochem. 2022; 171:429-441)解决了 FbTam41(一种来自真菌的功能直向同源物)与 CTP-Mg2+ 的晶体结构,成功地提供了 CDP-DAG 合成的详细分子观点。在这篇评论中,我们讨论了目前对 Tam41 介导的反应的理解。
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引用次数: 0
The action of coenzyme B12-dependent diol dehydratase on 3,3,3-trifluoro-1,2-propanediol results in elimination of all the fluorides with formation of acetaldehyde. 辅酶 B12 依赖性二元醇脱水酶对 3,3,3-三氟-1,2-丙二醇的作用会消除所有氟化物并形成乙醛。
IF 2.1 4区 生物学 Q4 BIOCHEMISTRY & MOLECULAR BIOLOGY Pub Date : 2024-09-03 DOI: 10.1093/jb/mvae047
Koichi Mori, Bernard T Golding, Tetsuo Toraya

3,3,3-Trifluoro-1,2-propanediol undergoes complete defluorination in two distinct steps: first, the conversion into 3,3,3-trifluoropropionaldehyde catalyzed by adenosylcobalamin (coenzyme B12)-dependent diol dehydratase; second, non-enzymatic elimination of all three fluorides from this aldehyde to afford malonic semialdehyde (3-oxopropanoic acid), which is decarboxylated to acetaldehyde. Diol dehydratase accepts 3,3,3-trifluoro-1,2-propanediol as a relatively poor substrate, albeit without significant mechanism-based inactivation of the enzyme during catalysis. Optical and electron paramagnetic resonance (EPR) spectra revealed the steady-state formation of cob(II)alamin and a substrate-derived intermediate organic radical (3,3,3-trifluoro-1,2-dihydroxyprop-1-yl). The coenzyme undergoes Co-C bond homolysis initiating a sequence of reaction by the generally accepted pathway via intermediate radicals. However, the greater steric size of trifluoromethyl and especially its negative impact on the stability of an adjacent radical centre compared to a methyl group has implications for the mechanism of the diol dehydratase reaction. Nevertheless, 3,3,3-trifluoropropionaldehyde is formed by the normal diol dehydratase pathway, but then undergoes non-enzymatic conversion into acetaldehyde, probably via 3,3-difluoropropenal and malonic semialdehyde.

3,3,3-三氟-1,2-丙二醇通过两个不同的步骤进行完全脱氟:首先,在依赖腺苷钴胺(辅酶 B12)的二元醇脱水酶催化下转化为 3,3,3-三氟丙醛;其次,从该醛中非酶消除所有三个氟化物,生成丙二酸半醛(3-氧代丙酸),然后脱羧为乙醛。二醇脱水酶接受 3,3,3-三氟-1,2-丙二醇作为相对较差的底物,尽管在催化过程中不会出现基于机理的酶失活现象。光学和 EPR 光谱显示,钴(II)氨基和底物衍生的中间有机自由基(3,3,3-三氟-1,2-二羟基丙-1-基)稳态形成。辅酶发生 Co-C 键均解,通过中间自由基以公认的途径启动一系列反应。然而,与甲基相比,三氟甲基的立体尺寸更大,尤其是对邻近自由基中心的稳定性有负面影响,这对二元醇脱水酶反应的机理产生了影响。不过,3,3,3-三氟丙醛是通过正常的二元醇脱水酶途径形成的,但随后可能通过 3,3-二氟丙烯醛和丙二醛半醛非酶转化为乙醛。
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引用次数: 0
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