RSC Chemical Biology最新文献

{"title":"P450 cyptide synthase MpoB catalyzes the cross-linking of the YPW motif on the precursor peptide.","authors":"Abujunaid Habib Khan, Jabal Rahmat Haedar, Vic Kiselov, Viktors Romanuks, Gints Smits, Stefano Donadio, Chin-Soon Phan","doi":"10.1039/d5cb00153f","DOIUrl":"https://doi.org/10.1039/d5cb00153f","url":null,"abstract":"<p><p>Cytochrome P450 enzymes in ribosomally synthesized and post-translationally modified peptides (RiPPs) catalyze C-C, C-N, or C-O cross-linking reactions in the biosynthesis of biaryl cyclophane natural products. Here, we manually identified 127 homologous P450s linked to putative precursor peptides containing the YPW motif. Through <i>in vivo</i> functional studies in <i>Escherichia coli</i>, the newly identified enzyme MpoB from <i>Micromonospora polyrhachis</i> DSM 45886 was found to catalyze the formation of a cross-link between Tyr-C3 and Trp-N1 at the YPW motif. This result provides an additional toolkit for cross-linked peptide modification.</p>","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":3.1,"publicationDate":"2025-07-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12288599/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144733731","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":"Regioselective rapid ene-type reaction (RRER) enables bioconjugation of histone serotonylation.","authors":"Jinghua Wu, Huapeng Li, Adam R Lovato, Andrew Symasek, Zeng Lin, Qingfei Zheng","doi":"10.1039/d5cb00159e","DOIUrl":"10.1039/d5cb00159e","url":null,"abstract":"<p><p>Triazolinedione (TAD) derivatives have been commonly utilized as protection and labeling reagents for indole and phenol moieties <i>via</i> a reversible ene-type reaction. Previous studies showed that the TAD probes could selectively modify tyrosine and tryptophan side-chains within proteins and peptides under distinct pH conditions. Here, we report a pH-controlled regioselective rapid ene-type reaction (RRER) between TAD and 5-hydroxyindole, where the modification occurs on the C4 position rather than the C3 of inactivated indole rings. Employing this unique reaction, we have performed the selective bioconjugation of serotonylation occurring on the fifth amino acid residue, glutamine, of histone H3 (H3Q5), which does not contain any tryptophan in its protein sequence. Finally, RRER was applied to determine the H3Q5 serotonylation levels in cultured cells and tissue samples, which served as a newly developed powerful tool for <i>in vitro</i> and <i>in vivo</i> histone monoaminylation analysis. Overall, our findings in this research expanded the chemical biology toolbox for investigating histone monoaminylation and facilitated the understandings of TAD-mediated ene-type reactions.</p>","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-07-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12282475/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144699844","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":"Chemoenzymatic synthesis of sialylated and fucosylated mucin analogs reveals glycan-dependent effects on protein conformation and degradation.","authors":"Amanda M Wood, Casia L Wardzala, Jessica R Kramer","doi":"10.1039/d5cb00111k","DOIUrl":"10.1039/d5cb00111k","url":null,"abstract":"<p><p>Mucin proteins are essential for life but are challenging to study due to their complex glycosylation patterns. Synthetic mimics have become vital tools for understanding and modulating the roles of mucins in human health and disease. These materials also have diverse biomedical applications as lubricants and anti-infectives, in vaccine formulations, and more. We developed a chemoenzymatic approach to prepare polypeptide-based synthetic mucins displaying a variety of glycans with native linkages and orientations. By combining the polymerization of glycosylated amino acid <i>N</i>-carboxyanhydrides with enzymatic sialylation and fucosylation, we produced a tunable panel of synthetic mucins. These polymers were recognized by natural glycan-binding and glycan-degrading enzymes, providing insights into the structural preferences of these proteins. Glycan- and linkage-dependent effects on proteolysis were observed. Further, investigation of the influence of glycans on peptide backbone secondary structure revealed that both sialylation and linkage at Ser <i>vs.</i> Thr have profound effects on hierarchical conformation. Overall, our methodology offers versatile tools for exploring the diverse glycobiology of mucins.</p>","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-07-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12266245/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144660696","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":"Random peptide mixtures of tryptophan and lysine suppress the aggregation of a cancer-related mutant of the Axin protein.","authors":"Tommaso Garfagnini, Zvi Hayouka, Assaf Friedler","doi":"10.1039/d5cb00141b","DOIUrl":"10.1039/d5cb00141b","url":null,"abstract":"<p><p>Aggregation of dysfunctional proteins can lead to a variety of diseases including cancer. We have previously developed chaperone-derived peptides that inhibit aggregation of the cancer-related L106R mutant of Axin RGS. Here we show that significantly improved inhibition was achieved using random peptide mixtures (RPMs) designed to mimic the chemical characteristics of the chaperone-like peptides. 20-mer RPMs of tryptophan and lysine suppressed aggregation of Axin RGS L106R with up to 50-fold improved activity compared to parent inhibitors. Conversely, peptides derived from the lead hotspot of Axin RGS aggregation that were designed to be specific, were unable to prevent aggregation of the protein. RPMs constitute the most efficient strategy to date to magnify peptide inhibitory activity against Axin RGS L106R aggregation, as they contain multiple active species and conformations that cover a larger inhibitory space and shield multiple hotspots at once. Our results demonstrate that the chemical composition of the peptide, and not the specific sequence, is the key factor for inhibitory activity.</p>","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-07-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12264706/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144660697","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":"Signals from the sea: the structural peculiarity of lipid A and weak immunostimulatory lipopolysaccharide from <i>Rheinheimera japonica</i>.","authors":"Stefania De Chiara, Francesca Olmeo, Emanuela Andretta, Luca De Simone Carone, Marcello Mercogliano, Vlada S Belova, Lyudmila A Romanenko, Maxim S Kokoulin, Alba Silipo, Antonio Molinaro, Flaviana Di Lorenzo","doi":"10.1039/d5cb00134j","DOIUrl":"10.1039/d5cb00134j","url":null,"abstract":"<p><p>Lipopolysaccharides (LPSs) isolated from marine bacteria represent a valuable resource for biomedical innovation. Here, we report the first structural elucidation of the lipid A moiety and a preliminary immunological assessment of the full LPS from the marine Gram-negative <i>Rheinheimera japonica</i> KMM 9513^T. Using MALDI-TOF mass spectrometry (MS) and tandem MS, we show that the lipid A from <i>R. japonica</i> KMM 9513^T exhibits a heterogeneous architecture, composed of <i>mono</i>- and <i>bis</i>-phosphorylated tetra- and penta-acylated species with variations in the acyl chain length, saturation, branching, and positional isomerism. Functionally, the full LPS was found to be immunologically silent toward TLR4-mediated NF-κB activation in HEK-Blue™ hTLR4 cells and triggered only modest, dose-dependent responses in differentiated human THP-1 macrophages. Strikingly, the <i>R. japonica</i> LPS was able to antagonize <i>E. coli</i> LPS-induced TLR4 activation, even at low doses. Overall, this study uncovers a structurally and functionally atypical marine LPS with a dual profile, inactive towards TLR4 yet capable of modulating LPS-induced signaling. These findings offer a promising basis to consider <i>R. japonica</i> LPS as a source of structural inspiration for the design of synthetic derivatives with controlled immunological properties.</p>","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-07-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12268950/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144676018","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":"Exploring the immunoproteasome's substrate preferences for improved hydrolysis and selectivity.","authors":"Christine S Muli, Cody A Loy, Darci J Trader","doi":"10.1039/d5cb00114e","DOIUrl":"10.1039/d5cb00114e","url":null,"abstract":"<p><p>The proteasome is an integral macromolecular machine responsible for regulated protein degradation, and its barrel-like core particle (CP) hydrolyzes protein substrates into peptide fragments. A proteasome isoform that is expressed under conditions of inflammation is known as the immunoproteasome (iCP), which incorporates different catalytic subunits of altered cleavage specificities from the standard proteasome (sCP). Probes and inhibitors have been generated to study iCP activity and for therapeutics, respectively; recently, the iCP has been harnessed as a prodrug enzyme to release bioactive compounds selectively into iCP-expressing cells. iCP-targeting probes, prodrugs, and inhibitors are based on peptide recognition sequences and their favorable interactions within the iCP's substrate channel. To better understand what unnatural substrates the iCP can recognize, we synthesized peptide-conjugated substrates and applied them to a liquid chromatography-mass spectrometry (LC-MS) method after incubation with purified human iCP. Structure-activity relationships of unnatural peptide-conjugated substrates revealed modifications that improved substrate selectively for the iCP by more than 3-fold compared to the original scaffold. As such, this report will be helpful to guide future iCP-targeting probes, prodrugs, and inhibitor design.</p>","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-07-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12235443/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144601775","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":"Structure-guided engineering of a polyphosphate kinase 2 class III from an <i>Erysipelotrichaceae</i> bacterium to produce base-modified purine nucleotides.","authors":"Rachel M Mitton-Fry, René Rasche, Ann-Marie Lawrence-Dörner, Jannik Eschenbach, Aileen Tekath, Andrea Rentmeister, Daniel Kümmel, Nicolas V Cornelissen","doi":"10.1039/d5cb00108k","DOIUrl":"10.1039/d5cb00108k","url":null,"abstract":"<p><p>Nucleobase-modified nucleoside-5'-triphosphates (NTPs) are important building blocks for the enzymatic synthesis of non-coding RNAs and mRNAs with improved properties. Chemical phosphorylation of base-modified nucleotides to NTPs remains challenging. Here, we report the enzymatic phosphorylation of purine-modified nucleoside-5'-monophosphates (NMPs) to the corresponding NTPs by the polyphosphate kinase 2 class III from an <i>Erysipelotrichaceae</i> bacterium (EbPPK2). The enzyme is highly promiscuous, accepting a range of NMPs with purine modifications. EbPPK2 efficiently catalyses the formation of the corresponding di-, tri- and tetraphosphates, typically with >70% conversion to the NTP. Slower conversion was observed for analogues with oxo- or thio-substitutions at the C6-position. To better understand nucleotide binding and catalysis, we determined the crystal structure of EbPPK2 at 1.7 Å resolution bound to a non-hydrolysable ATP analogue and polyphosphate. This enabled structure-guided design of EbPPK2 variants that efficiently convert GMP analogues, while retaining activity for AMP. Apart from being the preferred industrial-scale ATP recycling catalyst, EbPPK2 and variants bear potential to become the favoured enzyme family for purine-modified NTP production.</p>","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-07-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12257356/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144643776","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":"Correction: Turn-on fluorescent glucose transport bioprobe enables wash-free real-time monitoring of glucose uptake activity in live cells and small organisms.","authors":"Monica S Hensley, David Hutchings, Aldelrahman Ismail, Micaela Rayne Geborkoff, Thomas Werner, Marina Tanasova","doi":"10.1039/d5cb90030a","DOIUrl":"https://doi.org/10.1039/d5cb90030a","url":null,"abstract":"<p><p>[This corrects the article DOI: 10.1039/D4CB00239C.].</p>","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-07-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12226788/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144576597","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":"A dual-functional substrate for quantitation of substrate levels and GCase activity in living cells.","authors":"Ben Tiet, Sha Zhu, Xi Chen, Nadia Anastasi, Nicholas W See, Matthew C Deen, Eva Harde, David J Vocadlo","doi":"10.1039/d5cb00045a","DOIUrl":"10.1039/d5cb00045a","url":null,"abstract":"<p><p>Loss of function mutations in the gene <i>GBA1</i>, which encodes the lysosomal glycoside hydrolase β-glucocerebrosidase (GCase) cause Gaucher's disease (GD). Moreover, one mutant allele of <i>GBA1</i> is the most common genetic risk factor for the development of Parkinson's disease (PD). To gain a better understanding how these mutations drive development of PD and how GCase is regulated within cells, the field needs chemical reporters of GCase activity that can be used within living cells. Fluorogenic substrates are one method that can be used to quantify enzyme activities within cells yet existing substrates for GCase have limitations. In particular, the inability to monitor cellular uptake of substrate limits the ability to disentangle impairments in uptake of substrate from impairments in lysosomal GCase activity. Here we report on the preparation and biological characterisation of LysoRF-GBA - a new chemical tool which can be used to quantitatively measure both the cellular levels of intact substrate and lysosomal GCase activity within lysosomes. We demonstrate that, by using LysoRF-GBA, endogenous GCase activity can be measured within live neuroblastoma cells. The selectivity of this substrate for GCase, relative to other cellular enzymes, was validated by genetic and pharmacological perturbation of GCase. By using LysoRF-GBA and concomitantly monitoring levels of both cleaved product and intact substrate, we were able to measure GCase engagement with a known pharmacological chaperone and discriminate between pharmacological agents that affect GCase activity from those that impair endocytosis. Further, the ability to monitor intracellular levels of intact LysoRF-GBA also enabled us to measure its time dependent accumulation within cells, providing insight into when steady state levels of this substrate are reached. LysoRF-GBA therefore shows high potential to be exploited as a tool for the discovery of compounds that could beneficially modulate its activity for benefit in diseases including PD.</p>","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-07-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12223415/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144576596","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":"A biparatopic HER2-targeting ADC constructed <i>via</i> site-specific glycan conjugation exhibits superior stability, safety, and efficacy.","authors":"Qi Xue, Jianjian Peng, Wenying Dai, Qingsong Wu, Jinbiao Jiao, Yudi Hu, Wanxing Sha, Yang Yang, Wenhao Yu, Siyang Liu, Ting Xu, Jie P Li","doi":"10.1039/d5cb00096c","DOIUrl":"10.1039/d5cb00096c","url":null,"abstract":"<p><p>HER2 is overexpressed in approximately 15-20% of cancers and is associated with aggressive disease progression. We developed JSKN003, a bispecific HER2-targeted antibody-drug conjugate (ADC), through site-specific conjugation technology based on <i>N</i>-glycosylation engineering. JSKN003 maintains a biantennary glycan structure and exhibits superior structural homogeneity, optimized hydrophilicity, and reduced aggregation compared to conventional thiol-maleimide chemistry. In preclinical models JSKN003 demonstrated potent antitumor efficacy, inducing tumor regression in multiple HER2-expressing tumors, such as NCI-N87, BxPC-3, and PDX tumor models. Mechanistically, JSKN003 binds specifically to HER2, undergoes efficient internalization, and traffics to the lysosome, where the payload DXd is released, leading to DNA damage and apoptosis. JSKN003 retained its cytotoxic activity against trastuzumab-resistant cells, attributed to efficient payload delivery and blockade of downstream HER2 signaling pathways, demonstrating the potential to overcome clinical trastuzumab resistance. The safety profile of JSKN003 was evaluated in cynomolgus monkeys and was found to be acceptable, with no severe toxicities observed at therapeutic doses. JSKN003 demonstrated excellent antitumor activity and a favorable safety profile in clinical trials, highlighting its potential as a promising therapeutic option for patients with HER2-positive tumors. These findings suggest that JSKN003 could be a valuable therapeutic strategy with excellent efficacy and safety for HER2-expressing tumors in the clinical setting.</p>","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-06-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12188319/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144508786","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}