RSC Chemical Biology最新文献_第9页

Reverse transcription as key step in RNA in vitro evolution with unnatural base pairs† 反转录是具有非天然碱基对的 RNA 体外进化的关键步骤

IF 4.1

RSC Chemical Biology Pub Date : 2024-04-23 DOI: 10.1039/D4CB00084F

Eva S. Hoffmann, Mareike C. De Pascali, Lukas Neu, Christof Domnick, Alice Soldà and Stephanie Kath-Schorr

{"title":"Reverse transcription as key step in RNA in vitro evolution with unnatural base pairs†","authors":"Eva S. Hoffmann, Mareike C. De Pascali, Lukas Neu, Christof Domnick, Alice Soldà and Stephanie Kath-Schorr","doi":"10.1039/D4CB00084F","DOIUrl":"10.1039/D4CB00084F","url":null,"abstract":"Unnatural base pairs (UBPs) augment the chemical diversity of artificial nucleic acids and can thus enable the generation of new aptamers and catalytic nucleic acids by in vitro selection. However, owing to a lack of methodologies, the reverse transcription of UBPs, a key step in RNA aptamer selection, has not been sufficiently characterized. Here, we present a series of versatile assays to investigate the reverse transcription of the TPT3:NaM base pair as a representative for hydrophobic unnatural base pairs. We determine the fidelity and retention of the UBP for four different reverse transcriptases (RT) in the context of RNA in vitro evolution. The retention of the TPT3:NaM pair during the RNA in vitro selection process was investigated using a novel click-chemistry based electromobility shift assay. Real-time monitoring of reverse transcription kinetics revealed considerable differences in polymerase activity processing the TPT3:NaM base pair. Our findings identified SuperScript IV RT as the most efficient RT for processing the TPT3:NaM pair. Our approach can be applied universally to study newly developed UBPs, not only at the reverse transcription level, but also during PCR and in vitro transcription.","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" 6","pages":" 556-566"},"PeriodicalIF":4.1,"publicationDate":"2024-04-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://pubs.rsc.org/en/content/articlepdf/2024/cb/d4cb00084f?page=search","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140798084","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Why pyridoxal phosphate could be a functional predecessor of thiamine pyrophosphate and speculations on a primordial metabolism† 为什么磷酸吡哆醛可能是焦磷酸硫胺素的功能性前身，以及对原始新陈代谢的推测

IF 4.1

RSC Chemical Biology Pub Date : 2024-04-18 DOI: 10.1039/D4CB00016A

Andreas Kirschning

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Ancient and modern mechanisms compete in progesterone receptor activation† 黄体酮受体激活过程中的古今竞争机制

IF 4.1

RSC Chemical Biology Pub Date : 2024-04-15 DOI: 10.1039/D4CB00002A

Sabab Hasan Khan, Namita Dube, Nishanti Sudhakar, Olivia Fraser, Priscilla Villalona, Sean M. Braet, Stephanie Leedom, Erin R. Reilly, Jacob Sivak, Kenidee Crittenden and C. Denise Okafor

{"title":"Ancient and modern mechanisms compete in progesterone receptor activation†","authors":"Sabab Hasan Khan, Namita Dube, Nishanti Sudhakar, Olivia Fraser, Priscilla Villalona, Sean M. Braet, Stephanie Leedom, Erin R. Reilly, Jacob Sivak, Kenidee Crittenden and C. Denise Okafor","doi":"10.1039/D4CB00002A","DOIUrl":"10.1039/D4CB00002A","url":null,"abstract":"The progesterone receptor (PR) belongs to the steroid receptor family of ligand-regulated transcription factors, controlling genes important for development, metabolism, and reproduction. Understanding how diverse ligands bind and modulate PR activity will illuminate the design of ligands that control PR-driven signaling pathways. Here, we use molecular dynamics simulations to investigate how PR dynamics are altered by functionally diverse ligands. Using a library of 33 steroidal ligands that range from inactive to EC50 < 0.1 nM, we reveal an unexpected evolutionary basis for the wide gamut of activation. While other oxosteroid receptors employ an evolutionarily conserved mechanism dependent on a hydrogen bond between the receptor and ligand, extant PR has evolved a preference for activation that is not reliant on this polar interaction. We demonstrate that potent ligands utilize the modern PR mechanism while weaker ligands coopt the defunct ancestral mechanism by forming hydrogen bonds with Asn719. Based on their structures and dynamic signatures, ligands partition into four classes (inactive, weak, moderate and high potency) that interact distinctly with the PR binding pocket. Further, we use luciferase reporter assays and PR mutants to probe the roles of pocket residues in mediating distinct PR mechanisms. This combination of MD simulations and in vitro studies provide insight into how the evolutionary history of PR shapes its response to diverse ligands.","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" 6","pages":" 518-529"},"PeriodicalIF":4.1,"publicationDate":"2024-04-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://pubs.rsc.org/en/content/articlepdf/2024/cb/d4cb00002a?page=search","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140574798","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Reduction midpoint potential of a paradigm light–oxygen–voltage receptor and its modulation by methionine residues† 范式光氧电压受体的还原中点电位及其受蛋氨酸残基的调节

IF 4.1

RSC Chemical Biology Pub Date : 2024-04-09 DOI: 10.1039/D4CB00056K

Andrés García de Fuentes and Andreas Möglich

{"title":"Reduction midpoint potential of a paradigm light–oxygen–voltage receptor and its modulation by methionine residues†","authors":"Andrés García de Fuentes and Andreas Möglich","doi":"10.1039/D4CB00056K","DOIUrl":"10.1039/D4CB00056K","url":null,"abstract":"Light-dependent adaptations of organismal physiology, development, and behavior abound in nature and depend on sensory photoreceptors. As one class, light–oxygen–voltage (LOV) photoreceptors harness flavin-nucleotide chromophores to sense blue light. Photon absorption drives the LOV receptor to its signaling state, characterized by a metastable thioadduct between the flavin and a conserved cysteine residue. With this cysteine absent, LOV receptors instead undergo photoreduction to the flavin semiquinone which however can still elicit downstream physiological responses. Irrespective of the cysteine presence, the LOV photochemical response thus entails a formal reduction of the flavin. Against this backdrop, we here investigate the reduction midpoint potential E0 in the paradigmatic LOV2 domain from Avena sativa phototropin 1 (AsLOV2), and how it can be deliberately varied. Replacements of residues at different sites near the flavin by methionine consistently increase E0 from its value of around −280 mV by up to 40 mV. Moreover, methionine introduction invariably impairs photoactivation efficiency and thus renders the resultant AsLOV2 variants less light-sensitive. Although individual methionine substitutions also affect the stability of the signaling state and downstream allosteric responses, no clear-cut correlation with the redox properties emerges. With a reduction midpoint potential near −280 mV, AsLOV2 and, by inference, other LOV receptors may be partially reduced inside cells which directly affects their light responsiveness. The targeted modification of the chromophore environment, as presently demonstrated, may mitigate this effect and enables the design of LOV receptors with stratified redox sensitivities.","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" 6","pages":" 530-543"},"PeriodicalIF":4.1,"publicationDate":"2024-04-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://pubs.rsc.org/en/content/articlepdf/2024/cb/d4cb00056k?page=search","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140574655","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Advances in the joint profiling technologies of 5mC and 5hmC 5mC 和 5hmC 联合分析技术的进展

IF 4.1

RSC Chemical Biology Pub Date : 2024-04-05 DOI: 10.1039/D4CB00034J

Bo He, Haojun Yao and Chengqi Yi

引用次数: 0

Polyamines promote xenobiotic nucleic acid synthesis by modified thermophilic polymerase mutants† 多胺促进改良嗜热聚合酶突变体合成异种核酸

IF 4.1

RSC Chemical Biology Pub Date : 2024-04-04 DOI: 10.1039/D4CB00017J

Hidekazu Hoshino, Yuuya Kasahara and Satoshi Obika

{"title":"Polyamines promote xenobiotic nucleic acid synthesis by modified thermophilic polymerase mutants†","authors":"Hidekazu Hoshino, Yuuya Kasahara and Satoshi Obika","doi":"10.1039/D4CB00017J","DOIUrl":"10.1039/D4CB00017J","url":null,"abstract":"The enzymatic synthesis of xenobiotic nucleic acids (XNA), which are artificially sugar-modified nucleic acids, is essential for the preparation of XNA libraries. XNA libraries are used in the in vitro selection of XNA aptamers and enzymes (XNAzymes). Efficient enzymatic synthesis of various XNAs can enable the screening of high-quality XNA aptamers and XNAzymes by expanding the diversity of XNA libraries and adding a variety of properties to XNA aptamers and XNAzymes. However, XNAs that form unstable duplexes with DNA, such as arabino nucleic acid (ANA), may dissociate during enzyme synthesis at temperatures suitable for thermophilic polymerases. Thus, such XNAs are not efficiently synthesised by the thermophilic polymerase mutants at the end of the sequence. This undesirable bias reduces the possibility of generating high-quality XNA aptamers and XNAzymes. Here, we demonstrate that polyamine-induced DNA/ANA duplex stabilisation promotes ANA synthesis that is catalysed by thermophilic polymerase mutants. Several polyamines, including spermine, spermidine, cadaverine, and putrescine promote ANA synthesis. The negative effect of polyamines on the fidelity of ANA synthesis was negligible. We also showed that polyamines promote the synthesis of other XNAs, including 2′-amino-RNA/2′-fluoro-RNA mixture and 2′-O-methyl-RNA. In addition, we found that polyamine promotes DNA synthesis from the 2′-O-methyl-RNA template. Polyamines, with the use of thermophilic polymerase mutants, may allow further development of XNA aptamers and XNAzymes by promoting the transcription and reverse transcription of XNAs.","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" 5","pages":" 467-472"},"PeriodicalIF":4.1,"publicationDate":"2024-04-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://pubs.rsc.org/en/content/articlepdf/2024/cb/d4cb00017j?page=search","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140574529","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Prevention of amyloid β fibril deposition on the synaptic membrane in the precuneus by ganglioside nanocluster-targeting inhibitors† 神经节苷脂纳米簇靶向抑制剂可防止淀粉样β纤维沉积在楔前突触膜上

IF 4.1

RSC Chemical Biology Pub Date : 2024-04-04 DOI: 10.1039/D4CB00038B

Erika Miyamoto, Hideki Hayashi, Shigeo Murayama, Katsuhiko Yanagisawa, Toshinori Sato and Teruhiko Matsubara

{"title":"Prevention of amyloid β fibril deposition on the synaptic membrane in the precuneus by ganglioside nanocluster-targeting inhibitors†","authors":"Erika Miyamoto, Hideki Hayashi, Shigeo Murayama, Katsuhiko Yanagisawa, Toshinori Sato and Teruhiko Matsubara","doi":"10.1039/D4CB00038B","DOIUrl":"10.1039/D4CB00038B","url":null,"abstract":"Alzheimer's disease (AD), a progressive neurodegenerative condition, is one of the most common causes of dementia. Senile plaques, a hallmark of AD, are formed by the accumulation of amyloid β protein (Aβ), which starts to aggregate before the onset of the disease. Gangliosides, sialic acid-containing glycosphingolipids, play a key role in the formation of toxic Aβ aggregates. In membrane rafts, ganglioside-bound complexes (GAβ) act as nuclei for Aβ assembly, suggesting that GAβ is a promising target for AD therapy. The formation of GAβ-induced Aβ assemblies has been evaluated using reconstituted planar lipid membranes composed of synaptosomal plasma membrane (SPM) lipids extracted from human and mouse brains. Although the effects of gangliosides on Aβ accumulation in the precuneus have been established, effects on Aβ fibrils have not been determined. In this study, Aβ42 fibrils on reconstituted membranes composed of SPM lipids prepared from the precuneus cortex of human autopsied brains were evaluated by atomic force microscopy. In particular, Aβ42 accumulation, as well as the fibril number and size were higher for membranes with precuneus lipids than for membranes with calcarine cortex lipids. In addition, artificial peptide inhibitors targeting Aβ-sensitive ganglioside nanoclusters cleared Aβ assemblies on synaptic membranes in the brain, providing a novel therapeutic strategy for AD.","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" 5","pages":" 459-466"},"PeriodicalIF":4.1,"publicationDate":"2024-04-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://pubs.rsc.org/en/content/articlepdf/2024/cb/d4cb00038b?page=search","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140574642","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Site-specific RNA modification via initiation of in vitro transcription reactions with m6A and isomorphic emissive adenosine analogs† 通过 m6A 和同构发射型腺苷类似物启动体外转录反应对特定位点 RNA 进行修饰

IF 4.1

RSC Chemical Biology Pub Date : 2024-03-27 DOI: 10.1039/D4CB00045E

Deyuan Cong, Kfir B. Steinbuch, Ryosuke Koyama, Tyler V. Lam, Jamie Y. Lam and Yitzhak Tor

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Genome-wide mapping of G-quadruplex DNA: a step-by-step guide to select the most effective method G-quadruplex DNA 的全基因组图谱：选择最有效方法的分步指南。

IF 4.1

RSC Chemical Biology Pub Date : 2024-03-25 DOI: 10.1039/D4CB00023D

Silvia Galli, Gem Flint, Lucie Růžičková and Marco Di Antonio

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Introduction to ‘Medicinal Chemistry Small Molecule Probes’ 药物化学小分子探针 "简介