RSC Chemical Biology最新文献_第2页

Covalent functionalization of G protein-coupled receptors by small molecular probes.

IF 4.2

RSC Chemical Biology Pub Date : 2025-02-14 DOI: 10.1039/d4cb00294f

Bert L H Beerkens, Adriaan P IJzerman, Laura H Heitman, Daan van der Es

{"title":"Covalent functionalization of G protein-coupled receptors by small molecular probes.","authors":"Bert L H Beerkens, Adriaan P IJzerman, Laura H Heitman, Daan van der Es","doi":"10.1039/d4cb00294f","DOIUrl":"10.1039/d4cb00294f","url":null,"abstract":"Roughly one-third of all marketed drugs act by binding to one or more of the >800 human GPCRs, primarily through activation or inhibition via the orthosteric binding site. In addition, novel strategies to alter GPCR functioning are being developed, including allosteric, biased and covalently binding ligands. Molecular probes play an important role in verifying such drug molecules with new modes of action and providing information on all factors involved in GPCR signalling. Various types of molecular probes have been developed, ranging from small molecules to antibodies, each bearing its own advantages and disadvantages. In this mini-review, a closer look is taken at small molecular probes that functionalize GPCRs in a covalent manner, such as through the conjugation of reporter groups like fluorophores or biotin. Covalently bound reporter groups allow the investigation of GPCRs across an increasing range of biochemical assay types, yielding new insights into GPCR signalling pathways. Here, a broad range of recently developed 'functionalized covalent probes' is summarized. Furthermore, the use of these probes in biochemical assays and their applications in the field of GPCR research are discussed. Lastly, a view on possible future applications of these types of small molecular probes is provided.","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-02-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11827490/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143434001","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

Decoding allosteric landscapes: computational methodologies for enzyme modulation and drug discovery.

IF 4.2

RSC Chemical Biology Pub Date : 2025-02-14 DOI: 10.1039/d4cb00282b

Ruidi Zhu, Chengwei Wu, Jinyin Zha, Shaoyong Lu, Jian Zhang

{"title":"Decoding allosteric landscapes: computational methodologies for enzyme modulation and drug discovery.","authors":"Ruidi Zhu, Chengwei Wu, Jinyin Zha, Shaoyong Lu, Jian Zhang","doi":"10.1039/d4cb00282b","DOIUrl":"10.1039/d4cb00282b","url":null,"abstract":"Allosteric regulation is a fundamental mechanism in enzyme function, enabling dynamic modulation of activity through ligand binding at sites distal to the active site. Allosteric modulators have gained significant attention due to their unique advantages, including enhanced specificity, reduced off-target effects, and the potential for synergistic interaction with orthosteric agents. However, the inherent complexity of allosteric mechanisms has posed challenges to the systematic discovery and design of allosteric modulators. This review discusses recent advancements in computational methodologies for identifying and characterizing allosteric sites in enzymes, emphasizing techniques such as molecular dynamics (MD) simulations, enhanced sampling methods, normal mode analysis (NMA), evolutionary conservation analysis, and machine learning (ML) approaches. Advanced tools like PASSer, AlloReverse, and AlphaFold have further enhanced the understanding of allosteric mechanisms and facilitated the design of selective allosteric modulators. Case studies on enzymes such as Sirtuin 6 (SIRT6) and MAPK/ERK kinase (MEK) demonstrate the practical applications of these approaches in drug discovery. By integrating computational predictions with experimental validation, this review highlights the transformative potential of computational strategies in advancing allosteric drug discovery, offering innovative opportunities to regulate enzyme activity for therapeutic benefits.","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-02-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11836628/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143469649","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

The evolution and application of RNA-focused small molecule libraries.

IF 4.2

RSC Chemical Biology Pub Date : 2025-02-13 DOI: 10.1039/d4cb00272e

Amirhossein Taghavi, Noah A Springer, Patrick R A Zanon, Yanjun Li, Chenglong Li, Jessica L Childs-Disney, Matthew D Disney

引用次数: 0

Chemical approaches to explore ubiquitin-like proteins.

IF 4.2

RSC Chemical Biology Pub Date : 2025-02-12 DOI: 10.1039/d4cb00220b

Reem Mousa, Dana Shkolnik, Yam Alalouf, Ashraf Brik

{"title":"Chemical approaches to explore ubiquitin-like proteins.","authors":"Reem Mousa, Dana Shkolnik, Yam Alalouf, Ashraf Brik","doi":"10.1039/d4cb00220b","DOIUrl":"10.1039/d4cb00220b","url":null,"abstract":"Chemical protein synthesis has emerged as a powerful approach for producing ubiquitin (Ub) and ubiquitin-like modifiers (Ubls) in both their free and conjugated forms, particularly when recombinant or enzymatic strategies are challenging. By providing precise control over the assembly of Ub and Ubls, chemical synthesis enables the generation of complex constructs with site-specific modifications that facilitate detailed functional and structural studies. Ub and Ubls are central regulators of protein homeostasis, regulating a wide range of cellular processes such as cell cycle progression, transcription, DNA repair, and apoptosis. Ubls share an evolutionary link with Ub, resembling its structure and following a parallel conjugation pathway that results in a covalent isopeptide bond with their cellular substrates. Despite their structural similarities and sequence homology, Ub and Ubls exhibit distinct functional differences. Understanding Ubl biology is essential for unraveling how cells maintain their regulatory networks and how disruptions in these pathways contribute to various diseases. In this review, we highlight the chemical methodologies and strategies available for studying Ubls and advancing our comprehensive understanding of the Ubl system in health and disease.","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-02-12","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11817102/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143415684","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

Protein ligation for the assembly and study of nonribosomal peptide synthetase megaenzymes.

IF 4.2

RSC Chemical Biology Pub Date : 2025-02-07 DOI: 10.1039/d4cb00306c

Angelos Pistofidis, T Martin Schmeing

{"title":"Protein ligation for the assembly and study of nonribosomal peptide synthetase megaenzymes.","authors":"Angelos Pistofidis, T Martin Schmeing","doi":"10.1039/d4cb00306c","DOIUrl":"10.1039/d4cb00306c","url":null,"abstract":"Nonribosomal peptide synthetases (NRPSs) are biosynthetic enzymes found in bacteria and fungi, that synthesize a plethora of pharmaceutically relevant compounds. NRPSs consist of repeating sets of functional domains called modules, and each module is responsible for the incorporation of a single amino acid to the growing peptidyl intermediate. The synthetic logic of an NRPS resembles an assembly line, with growing biosynthesis intermediates covalently attached to the prosthetic 4'-phosphopantetheine (ppant) moieties of T (thiolation or transfer) domains for shuttling within and between modules. Therefore, NRPSs must have each T domain phosphopantetheinylated to be functional, and host organisms encode ppant transferases that affix ppant to T domains. Ppant transferases can be promiscuous with respect to the T domain substrate and with respect to chemical modifications of the ppant thiol, which has been a useful characteristic for study of megaenzymes and other systems. However, defined studies of multimodular megaenzymes, where different analogs are required to be affixed to different T domains within the same multimodular protein, are hindered by this promiscuity. Study of NRPS peptide bond formation, for which two T domains simultaneously deliver substrates to the condensation domain, is a prime example where one would want two T domains bearing different acyl/peptidyl groups. Here, we report a strategy where two NRPS modules that are normally part of the same protein are expressed as separate constructs, modified separately with different acyl-ppants, and then ligated together by sortase A of Staphylococcus aureus or asparaginyl endopeptidase 1 of Oldenlandia affinis (OaAEP1). We assessed various reaction conditions to optimize the ligation reactions and maximize the yield of the complex of interest. Finally, we apply this method in large scale and show it allows the complex built by OaAEP1-mediated ligation to be characterized by X-ray crystallography.","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-02-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11824870/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143434022","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

Cryo-electron microscopy reveals a single domain antibody with a unique binding epitope on fibroblast activation protein alpha.

IF 4.2

RSC Chemical Biology Pub Date : 2025-02-06 DOI: 10.1039/d4cb00267a

Zhen Xu, Akesh Sinha, Darpan N Pandya, Nicholas J Schnicker, Thaddeus J Wadas

引用次数: 0

Development of a near-infrared fluorescent probe for the selective detection of severe hypoxia†

IF 4.2

RSC Chemical Biology Pub Date : 2025-02-04 DOI: 10.1039/D4CB00243A

Takafumi Kasai, Kyohhei Fujita, Toru Komatsu, Tasuku Ueno, Ryosuke Kojima, Kenjiro Hanaoka and Yasuteru Urano

引用次数: 0

An activation-based high throughput screen identifies caspase-10 inhibitors.

IF 4.2

RSC Chemical Biology Pub Date : 2025-02-04 DOI: 10.1039/d5cb00017c

José O Castellón, Constance Yuen, Brandon Han, Katrina H Andrews, Samuel Ofori, Ashley R Julio, Lisa M Boatner, Maria F Palafox, Nithesh Perumal, Robert Damoiseaux, Keriann M Backus

{"title":"An activation-based high throughput screen identifies caspase-10 inhibitors.","authors":"José O Castellón, Constance Yuen, Brandon Han, Katrina H Andrews, Samuel Ofori, Ashley R Julio, Lisa M Boatner, Maria F Palafox, Nithesh Perumal, Robert Damoiseaux, Keriann M Backus","doi":"10.1039/d5cb00017c","DOIUrl":"10.1039/d5cb00017c","url":null,"abstract":"Caspases are a family of highly homologous cysteine proteases that play critical roles in inflammation and apoptosis. Small molecule inhibitors are useful tools for studying caspase biology, complementary to genetic approaches. However, achieving inhibitor selectivity for individual members of this highly homologous enzyme family remains a major challenge in developing such tool compounds. Prior studies have revealed that one strategy to tackle this selectivity gap is to target the precursor or zymogen forms of individual caspases, which share reduced structural homology when compared to active proteases. To establish a screening assay that favors the discovery of zymogen-directed caspase-10 selective inhibitors, we engineered a low-background and high-activity tobacco etch virus (TEV)-activated caspase-10 protein. We then subjected this turn-on protease to a high-throughput screen of approximately 100 000 compounds, with an average Z' value of 0.58 across all plates analyzed. Counter screening, including against TEV protease, delineated bona fide procaspase-10 inhibitors. Confirmatory studies identified a class of thiadiazine-containing compounds that undergo isomerization and oxidation to generate cysteine-reactive compounds with caspase-10 inhibitory activity. In parallel, mode-of-action studies revealed that pifithrin-μ (PFTμ), a reported TP53 inhibitor, also functions as a promiscuous caspase inhibitor. Both inhibitor classes showed preferential zymogen inhibition. Given the generalized utility of activation assays, we expect our screening platform to have widespread applications in identifying state-specific protease inhibitors.","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/PMC11854450/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143516947","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

Photothermal imaging of cellular responses to glucose deprivation.

IF 4.2

RSC Chemical Biology Pub Date : 2025-01-31 DOI: 10.1039/d4cb00269e

Jun Miyazaki, Ryotaro Wagatsuma, Koji Okamoto

{"title":"Photothermal imaging of cellular responses to glucose deprivation.","authors":"Jun Miyazaki, Ryotaro Wagatsuma, Koji Okamoto","doi":"10.1039/d4cb00269e","DOIUrl":"10.1039/d4cb00269e","url":null,"abstract":"In solid tumours, cancer cells modify their metabolic processes to endure environments with nutrient and oxygen scarcity due to inadequate blood flow. A thorough understanding of this adaptive mechanism, which requires reliable microscopic techniques, is crucial for developing effective cancer treatments. In the present study, we used multi-wavelength photothermal (PT) microscopy to visualise the cellular response to glucose deprivation in living cells derived from cervical cancer. We found increased mitochondrial PT signal intensity under glucose deprivation conditions, which is indicative of a correlation between mitochondrial crista density and PT signal intensity. Furthermore, PT microscopy revealed that the activity of the autophagy-lysosome system can be evaluated by detecting substances accumulated in lysosomes. Using this method, we confirmed that ferritin and denatured proteins from the endoplasmic reticulum were present within the lysosomes. The detectability of these substances using PT microscopy at visible wavelengths indicated the presence of iron ions. This method does not require labeling of molecules and provides reliable information and detailed insights into the cellular responses associated with the adaptation of cancer cell metabolism to nutrient stress conditions.","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-01-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11801213/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143383605","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

Regulating ferredoxin electron transfer using nanobody and antigen interactions.

IF 4.2

RSC Chemical Biology Pub Date : 2025-01-31 DOI: 10.1039/d4cb00257a

Albert Truong, Jonathan J Silberg

{"title":"Regulating ferredoxin electron transfer using nanobody and antigen interactions.","authors":"Albert Truong, Jonathan J Silberg","doi":"10.1039/d4cb00257a","DOIUrl":"10.1039/d4cb00257a","url":null,"abstract":"Fission and fusion can be used to generate new regulatory functions in proteins. This approach has been used to create ferredoxins (Fd) whose cellular electron transfer is dependent upon small molecule binding. To investigate whether Fd fragments can be used to monitor macromolecular binding reactions, we investigated the effects of fusing fragments of Mastigocladus laminosus Fd to single domain antibodies, also known as nanobodies, and their protein antigens. When Fd fragments arising from fission were fused to green fluorescent protein (GFP) and three different anti-GFP nanobodies, split proteins were identified that supported Fd-mediated electron transfer from Fd-NADP reductase (FNR) to sulfite reductase (SIR) in Escherichia coli. However, the order of nanobody and antigen fusion to the Fd fragments affected cellular electron transfer. Insertion of these anti-GFP nanobodies within Fd had differing effects on electron transfer. One domain-insertion variant was unable to support cellular electron transfer unless it was coexpressed with GFP, while others supported electron transfer in the absence of GFP. These findings show how Fds can be engineered so that their electron transfer is regulated by macromolecules, and they reveal the importance of exploring different nanobody homologs and fusion strategies when engineering biomolecular switches.","PeriodicalId":40691,"journal":{"name":"RSC Chemical Biology","volume":" ","pages":""},"PeriodicalIF":4.2,"publicationDate":"2025-01-31","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11886610/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143587580","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