RSC Chemical Biology最新文献

{"title":"Development of a near-infrared fluorescent probe for the selective detection of severe hypoxia.","authors":"Takafumi Kasai, Kyohhei Fujita, Toru Komatsu, Tasuku Ueno, Ryosuke Kojima, Kenjiro Hanaoka, Yasuteru Urano","doi":"10.1039/d4cb00243a","DOIUrl":"10.1039/d4cb00243a","url":null,"abstract":"<p><p>Severely hypoxic environments with oxygen concentrations around 1% are often found in serious diseases such as ischemia and cancer. However, existing near-infrared (NIR) fluorescent probes that can visualize hypoxia are also activated in mildly hypoxic environments (around 5% oxygen). Here, in order to selectively detect severe hypoxia, we used julolidine-based SiR (JSiR) as a NIR fluorophore and developed T-azoJSiR640 as a fluorescent probe. T-azoJSiR640 was able to detect severe hypoxia (around 1% oxygen concentration or less) in live cell imaging. Furthermore, the ischemic liver in a portal-vein-ligated mouse model was successfully visualized <i>in vivo</i>.</p>","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-02-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11791654/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143256437","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}

{"title":"Site-specific <i>in vivo</i> protein SUMOylation <i>via</i> translational incorporation of a proximity-reactive pyrrolysine analogue.","authors":"Yuk Hei Chan, Marianne M Lee, Michael K Chan","doi":"10.1039/d4cb00135d","DOIUrl":"10.1039/d4cb00135d","url":null,"abstract":"<p><p>Here, we present a novel strategy that integrates genetic-code expansion and proximity-induced crosslinking to achieve site-specific <i>in vivo</i> SUMOylation. This approach involves incorporating the unnatural amino acid 2-chloroacetyl-<i>N</i>ε-lysine (ClAcK) into the target protein using MmFAcKRS1, a previously reported pyrrolysyl-tRNA synthetase mutant that we have repurposed for ClAcK incorporation. Once incorporated, ClAcK can be specifically targeted to react with a cysteine engineered at the C-terminus of SUMO variants leading to a chemically SUMOylated protein. This reaction is proximity-induced, and preferentially promoted when the two reactive groups are in close spatial proximity. We therefore leverage the natural affinity of SUMO for SUMO-interacting motifs (SIMs) on target proteins to generate the targeted SUMO conjugation. Using this approach, site-specific SUMO-conjugates have been produced for two distinct proteins in cells, thus demonstrating its potential as a strategy for helping to dissect the role of SUMOylation in its native cellular context.</p>","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-01-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11791516/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143256544","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}

{"title":"Biochemical characterization and discovery of inhibitors for <i>Pf</i>Sir2A: new tricks for an old enzyme.","authors":"Dickson Donu, Emily Boyle, Alyson Curry, Yana Cen","doi":"10.1039/d4cb00206g","DOIUrl":"10.1039/d4cb00206g","url":null,"abstract":"<p><p>The Sir2 enzyme from <i>Plasmodium falciparum</i> (<i>Pf</i>Sir2A) is essential for the antigenic variation of this parasite, and its inhibition is expected to have therapeutic effects for malaria. Selective <i>Pf</i>Sir2A inhibitors are not available yet, partially due to the fact that this enzyme demonstrates extremely weak <i>in vitro</i> deacetylase activity, making the characterization of its inhibitors rather challenging. In the current study, we report the biochemical characterization and inhibitor discovery for this enzyme. <i>Pf</i>Sir2A exhibits greater enzymatic activity in the presence of DNA for both the peptide and histone protein substrates, suggesting that nucleosomes may be the real substrates of this enzyme. Indeed, it demonstrates robust deacetylase activity against nucleosome substrates, stemming primarily from the tight binding interactions with the nucleosome. In addition to DNA/nucleosome, free fatty acids (FFAs) are also identified as endogenous <i>Pf</i>Sir2A regulators. Myristic acid, a biologically relevant FFA, shows differential regulation of the two distinct activities of <i>Pf</i>Sir2A: activates deacetylation, but inhibits defatty-acylation. The structural basis of this differential regulation was further explored. Moreover, synthetic small molecule inhibitors of <i>Pf</i>Sir2A were discovered through the screening of a library of human sirtuin regulators. The mechanism of inhibition of the lead compounds were investigated. Collectively, the mechanistic insights and inhibitors described in this study will facilitate the future development of small molecule <i>Pf</i>Sir2A inhibitors as antimalarial agents.</p>","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-01-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11784564/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143081504","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}

{"title":"Raman signatures of type A and B influenza viruses: molecular origin of the “catch and kill” inactivation mechanism mediated by micrometric silicon nitride powder†","authors":"Giuseppe Pezzotti, Yoshiki Yasukochi, Eriko Ohgitani, Maiko Nakashio, Masaharu Shin-Ya, Tetsuya Adachi, Toshiro Yamamoto, Saki Ikegami, Wenliang Zhu, Koichiro Higasa, Kazu Okuma and Osam Mazda","doi":"10.1039/D4CB00237G","DOIUrl":"10.1039/D4CB00237G","url":null,"abstract":"<p >A multiomic study of the structural characteristics of type A and B influenza viruses by means of highly spectrally resolved Raman spectroscopy is presented. Three virus strains, A H1N1, A H3N2, and B98, were selected because of their known structural variety and because they have co-circulated with variable relative prevalence within the human population since the re-emergence of the H1N1 subtype in 1977. Raman signatures of protein side chains tyrosine, tryptophan, and histidine revealed unequivocal and consistent differences for pH characteristics at the virion surface, while different conformations of two C–S bond configurations in <em>gauche</em> and <em>trans</em> methionine rotamers provided distinct low-wavenumber fingerprints for different virus lineages/subtypes. Short-term exposure to a few percent fraction of silicon nitride (Si<small>₃</small>N<small>₄</small>) micrometric powder in an aqueous environment completely inactivated the influenza virions, independent of lineage/subtype dependent characteristics. The molecular-scale details of the inactivation process were studied by Raman spectroscopy and interpreted in terms of a “<em>catch and kill</em>” mechanism, in which the hydrolyzing ceramic surface first attracts virions with high efficiency through electrochemical interactions (mimicking cellular sialic acid) and then “poisons” the viruses by local hydrolytic elution of ammonia and nitrogen radicals. The latter event causes severe damage to the virions’ structures, including structural degradation of RNA purines, rotameric scrambling of methionine residues, formation of sulfhydryl and ionized carboxyl groups, and deprotonation/torsional deformation of tyrosine, tryptophan, and histidine residues. This study confirmed the antiviral effectiveness of Si<small>₃</small>N<small>₄</small> powder, which is safe to the human body and simply activated by water molecules. Raman spectroscopy was confirmed as a powerful tool in molecular virology, complementary to genomics and unique in providing direct information on virus structures at the molecular scale.</p>","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" 2","pages":" 182-208"},"PeriodicalIF":4.2,"publicationDate":"2025-01-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11751685/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143029882","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}

{"title":"RPRD1B's direct interaction with phosphorylated RNA polymerase II regulates polyadenylation of cell cycle genes and drives cancer progression.","authors":"Rosamaria Y Moreno, Svetlana B Panina, Y Jessie Zhang","doi":"10.1039/d4cb00212a","DOIUrl":"10.1039/d4cb00212a","url":null,"abstract":"<p><p>RNA polymerase II (Pol II) regulates eukaryotic gene expression through dynamic phosphorylation of its C-terminal domain (CTD). Phosphorylation at Ser2 and Thr4 on the CTD is crucial for RNA 3' end processing and facilitating the recruitment of cleavage and termination factors. However, the transcriptional roles of most CTD-binding proteins remain poorly understood. In this study, we focus on RPRD1B, a transcriptional regulator that interacts with the phosphorylated CTD and has been implicated in various cancers. We investigated its molecular mechanism during transcription and found that RPRD1B modulates alternative polyadenylation of cell growth transcripts by directly interacting with the CTD. RPRD1B is recruited to transcribing Pol II near the 3' end of the transcript, specifically in response to Ser2 and Thr4 phosphorylation, but only after flanking Ser5 phosphorylation is removed. Transcriptomic analysis of RPRD1B knockdown cells revealed its role in cell proliferation <i>via</i> termination of the key cell growth genes at upstream polyadenylation sites, leading to the production of tumor suppressor transcripts that lack AU-rich elements (AREs) with increased mRNA stability. Overall, our study uncovers previously unrecognized connections between the Pol II CTD and CID, highlighting their influence on 3' end processing and their contribution to abnormal cell growth in cancer.</p>","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-01-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11775580/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143068394","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}

{"title":"Novel photocrosslinking chemical probes utilized for high-resolution spatial transcriptomics.","authors":"Leslie Spitalny, Natalie Falco, Whitney England, Tyler Allred, Robert C Spitale","doi":"10.1039/d4cb00262h","DOIUrl":"10.1039/d4cb00262h","url":null,"abstract":"<p><p>The architecture of cells and the tissue they form within multicellular organisms are highly complex and dynamic. Cells optimize their function within tissue microenvironments by expressing specific subsets of RNAs. Advances in cell tagging methods enable spatial understanding of RNA expression when merged with transcriptomics. However, these techniques are currently limited by the spatial resolution of the tagging, the number of RNAs that can be sequenced, and multiplexing to isolate spatially-distinct cells within the same tissue landscape. To address these limitations, we developed CrossSeq, which employs photocrosslinking fluorescent probes and confocal microscopy activation to demarcate user-defined regions of interest on fixed cells for multiplexed spatial transcriptomic analysis. We investigate phenyl azide and diazirine crosslinking scaffolds and define their photoactivity profiles. We then deploy the aryl azide scaffold with three fluorophores for multiplexing on glyoxal fixed cells and analyze the defined populations using flow cytometry. Finally, we apply CrossSeq to investigate an <i>in vitro</i> MDA-MB-231-LM2 metastatic cancer migration model to evaluate changes in gene expression at the migratory cell front <i>versus</i> the exterior population. We anticipate this new technology will be a valuable tool addition as it will enable easier access to spatial transcriptomic analysis for the scientific community using conventional microscopy and analysis techniques.</p>","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-01-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11748054/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143025085","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}

{"title":"Bioinformatic and biochemical analysis uncovers novel activity in the 2-ER subfamily of OYEs.","authors":"Tamra C Blue-Lahom, Stacey K Jones, Katherine M Davis","doi":"10.1039/d4cb00289j","DOIUrl":"10.1039/d4cb00289j","url":null,"abstract":"<p><p>Members of the old yellow enzyme (OYE) family utilize a flavin mononucleotide cofactor to catalyze the asymmetric reduction of activated alkenes. The 2-enoate reductase (2-ER) subfamily are of particular industrial relevance as they can reduce α/β alkenes near electron-withdrawing groups. While the broader OYE family is being extensively explored for biocatalytic applications, oxygen sensitivity and poor expression yields associated with the presence of an Fe/S cluster in 2-ERs have hampered their characterization. Herein, we explore the use of pseudo-anaerobic preparation as a route to more widespread study of these enzymes and apply bioinformatics approaches to identify a subset of 2-ERs containing unusual mutations in both a key catalytic residue and the Fe/S cluster-binding motif. Biochemical analysis of a representative member from <i>Burkholderia insecticola</i> (OYE<i>Bi</i>) reveals novel <i>N</i>-methyl-proline demethylation activity, which we hypothesize may play a role in osmotic stress regulation based on genomic neighborhood analysis.</p>","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-01-20","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11759058/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143048385","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}

{"title":"Navigating the dichotomy of reactive oxygen, nitrogen, and sulfur species: detection strategies and therapeutic interventions.","authors":"Prayasee Baruah, Dikshaa Padhi, Hariharan Moorthy, Madhu Ramesh, Thimmaiah Govindaraju","doi":"10.1039/d5cb00006h","DOIUrl":"10.1039/d5cb00006h","url":null,"abstract":"<p><p>Reactive oxygen, nitrogen and sulfur species (RONSS) collectively encompasses a variety of energetically dynamic entities that emerge as inherent characteristics of aerobic life. This broad category includes reactive oxygen species (ROS), reactive nitrogen species (RNS), and reactive sulfur species (RSS). A conundrum arises from the indispensable role of RONSS in redox signalling, while its overproduction in the mitochondria poses deleterious effects. This imbalance leads to biomolecular damage and contributes to neurodegenerative disorders, cancer, cardiovascular diseases and inflammation. Notably, the differential roles of RONSS across various diseases can be strategically exploited for therapeutic interventions. Timely, precise, and sensitive detection methods are indispensable for elucidating the spatiotemporal dynamics of RONSS and evaluating disease pathogenesis and progression. By monitoring RONSS levels, we can discern early markers of disease onset, enabling proactive intervention strategies for effective disease management. Therapeutic interventions targeting oxidative/nitrosative stress in disease pathologies have proven to be effective treatment routes in the mitigation of different diseases. This review aims to offer a comprehensive overview of the functional implications and delicate balance of RONSS in disease conditions, and advances made in detection strategies over the years while offering therapeutic strategies to tackle their adverse effects. A special emphasis is focussed on neurodegenerative disorders and cancer with case studies using RONSS-targeted chemical probes and prodrugs.</p>","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-01-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11770382/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143060991","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}

{"title":"Siderophore-based targeted antibody recruitment for promoting immune responses towards Gram-negative pathogens.","authors":"Seungwoo Kim, Ho-Sung Park, Do Young Kim, Hyunhi Joh, Jiseok Oh, Dong Ho Kim, Min Ju Kang, Chul Hee Choi, Hak Joong Kim","doi":"10.1039/d4cb00293h","DOIUrl":"10.1039/d4cb00293h","url":null,"abstract":"<p><p>Antibody-recruiting molecules (ARMs) have emerged as a promising strategy for enhancing immune responses against pathogens and cancer cells. In this study, we developed a novel class of antibacterial ARMs utilizing siderophores, small iron-chelating compounds, as targeting motifs. Siderophores naturally exhibit high specificity for bacterial pathogens due to their role in iron acquisition, making them ideal candidates for selective targeting. We identified a potent ARM, GNP3, comprising MECAM, a siderophore mimetic, and 2,4-dinitrophenyl (DNP), a motif recognized by endogenous antibodies, connected <i>via</i> a flexible linker. GNP3 binds simultaneously to both anti-DNP antibody and the siderophore receptor, FepA, facilitating the targeted deposition of antibodies on the surface of FepA-expressing bacterial cells, such as <i>Escherichia coli</i> and <i>Pseudomonas aeruginosa</i>. This GNP3-induced opsonization promoted robust immune responses, including complement-dependent cytotoxicity (CDC) in the presence of serum and macrophage-mediated phagocytosis. Moreover, GNP3 effectively triggered CDC activity against serum-resistant uropathogenic <i>E. coli</i>. The results suggest that siderophore-based ARMs, by harnessing the immune defense system, represent a promising complementary approach to traditional antibiotics for overcoming recalcitrant bacterial infections.</p>","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-01-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11740091/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143013447","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}

{"title":"Raman active diyne-girder conformationally constrained p53 stapled peptides bind to MDM2 for visualisation without fluorophores.","authors":"Danielle C Morgan, Laura McDougall, Astrid Knuhtsen, Lori Buetow, Craig F Steven, Oscar A Shepperson, Danny T Huang, Alison N Hulme, Andrew G Jamieson","doi":"10.1039/d4cb00288a","DOIUrl":"10.1039/d4cb00288a","url":null,"abstract":"<p><p>Peptide stapling is an effective strategy to stabilise α-helical peptides, enhancing their bioactive conformation and improving physiochemical properties. In this study, we apply our novel diyne-girder stapling approach to the MDM2/MDMX α-helical binding region of the p53 transactivation domain. By incorporation of an unnatural amino acid to create an optimal <i>i</i>, <i>i</i> + 7 bridge length, we developed a highly α-helical stapled peptide, 4, confirmed <i>via</i> circular dichroism. This diyne-girder-stapled peptide demonstrated enhanced helicity and nanomolar binding affinity for MDM2, as assessed by fluorescence polarisation. Crucially, peptide 4 exhibited strong selectivity for MDM2, with approximately 100-fold reduced affinity for MDMX. Molecular modeling and docking studies suggested that this selectivity arose from diminished hydrophobic interactions at the MDMX binding site, driven by the diyne-girder's constrained geometry. The use of the diyne-girder, a unique feature amongst stapled peptide analogues, for cellular visualisation using Raman spectroscopy in the \"cell-silent\" region was explored. This capability potentially offers a novel method for tracking stapled peptides in biological systems without the need for large fluorophores. Overall, peptide 4 represents a promising tool for probing MDM2 activity and a valuable addition to the arsenal of peptide-based therapeutic strategies.</p>","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-01-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11741006/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143013442","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}