ACS Chemical Biology最新文献

Probing the Signal Transduction Mechanism of the Light-Activated Adenylate Cyclase OaPAC Using Unnatural Amino Acid Mutagenesis.

IF 3.5 2区生物学

ACS Chemical Biology Pub Date : 2025-01-22 DOI: 10.1021/acschembio.4c00627

Samruddhi S Jewlikar, Jinnette Tolentino Collado, Madeeha I Ali, Aya Sabbah, YongLe He, James N Iuliano, Christopher R Hall, Katrin Adamczyk, Gregory M Greetham, Andras Lukacs, Stephen R Meech, Peter J Tonge

{"title":"Probing the Signal Transduction Mechanism of the Light-Activated Adenylate Cyclase OaPAC Using Unnatural Amino Acid Mutagenesis.","authors":"Samruddhi S Jewlikar, Jinnette Tolentino Collado, Madeeha I Ali, Aya Sabbah, YongLe He, James N Iuliano, Christopher R Hall, Katrin Adamczyk, Gregory M Greetham, Andras Lukacs, Stephen R Meech, Peter J Tonge","doi":"10.1021/acschembio.4c00627","DOIUrl":"https://doi.org/10.1021/acschembio.4c00627","url":null,"abstract":"OaPAC, the photoactivated adenylyl cyclase from Oscillatoria acuminata, is composed of a blue light using FAD (BLUF) domain fused to an adenylate cyclase (AC) domain. Since both the BLUF and AC domains are part of the same protein, OaPAC is a model for understanding how the ultrafast modulation of the chromophore binding pocket caused by photoexcitation results in the activation of the output domain on the μs-s time scale. In the present work, we use unnatural amino acid mutagenesis to identify specific sites in the protein that are involved in transducing the signal from the FAD binding site to the ATP binding site. To provide insight into site-specific structural dynamics, we replaced W90 which is close to the chromophore pocket, F103 which interacts with W90 across the dimer interface, and F180 in the central core of the AC domain, with the infrared probe azido-Phe (AzPhe). Using ultrafast IR, we show that AzPhe at position 90 responds on multiple time scales following photoexcitation. In contrast, the light minus dark IR spectrum of AzPhe103 shows only a minor perturbation in environment between the dark and light states, while replacement of F180 with AzPhe resulted in a protein with no catalytic activity. We also replaced Y125, which hydrogen bonds with N256 across the dimer interface, with fluoro-Tyr residues. All the fluoro-Tyr substituted proteins retained the light-induced red shift in the flavin absorption spectrum; however, only the 3-FY125 OaPAC retained photoinduced catalytic activity. The loss of activity in 3,5-F2Y125 and 2,3,5-F3Y125 OaPAC, which potentially increase the acidity of the Y125 phenol by more than 1000-fold, suggests that deprotonation of Y125 disrupts the signal transduction pathway from the BLUF to the AC domain.","PeriodicalId":11,"journal":{"name":"ACS Chemical Biology","volume":" ","pages":""},"PeriodicalIF":3.5,"publicationDate":"2025-01-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143021284","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Tools for Intersectional Optical and Chemical Tagging on Cell Surfaces. 细胞表面交叉光学和化学标记工具。

IF 3.5 2区生物学

ACS Chemical Biology Pub Date : 2025-01-21 DOI: 10.1021/acschembio.4c00756

Sarah Innes-Gold, Hanzeng Cheng, Luping Liu, Adam E Cohen

引用次数: 0

Understanding the Glycosylation Pathways Involved in the Biosynthesis of the Sulfated Glycan Ligands for Siglecs. 糖基化途径在Siglecs硫酸化聚糖配体生物合成中的作用。

IF 3.5 2区生物学

ACS Chemical Biology Pub Date : 2025-01-21 DOI: 10.1021/acschembio.4c00677

Jaesoo Jung, Edward N Schmidt, Hua-Chien Chang, Zeinab Jame-Chenarboo, Jhon R Enterina, Kelli A McCord, Taylor E Gray, Lauren Kageler, Chris D St Laurent, Chao Wang, Ryan A Flynn, Peng Wu, Kay-Hooi Khoo, Matthew S Macauley

{"title":"Understanding the Glycosylation Pathways Involved in the Biosynthesis of the Sulfated Glycan Ligands for Siglecs.","authors":"Jaesoo Jung, Edward N Schmidt, Hua-Chien Chang, Zeinab Jame-Chenarboo, Jhon R Enterina, Kelli A McCord, Taylor E Gray, Lauren Kageler, Chris D St Laurent, Chao Wang, Ryan A Flynn, Peng Wu, Kay-Hooi Khoo, Matthew S Macauley","doi":"10.1021/acschembio.4c00677","DOIUrl":"https://doi.org/10.1021/acschembio.4c00677","url":null,"abstract":"Carbohydrate sulfation plays a pivotal role in modulating the strength of Siglec-glycan interactions. Recently, new aspects of Siglec binding to sulfated cell surface carbohydrates have been discovered, but the class of glycan presenting these sulfated Siglec ligands has not been fully elucidated. In this study, the contribution of different classes of glycans to cis and trans Siglec ligands was investigated within cells expressing the carbohydrate sulfotransferase 1 (CHST1) or CHST2. For some Siglecs, the glycan class mediating binding was clear, such as O-glycans for Siglec-7 and N-glycans for Siglec-2 and Siglec-9. Both N-glycans and mucin-type O-glycans contributed to ligands for Siglec-3, -5, -8, and -15. However, significant levels of Siglec-3 and -8 ligands remained in CHST1-expressing cells lacking complex N-glycans and mucin-type O-glycans. A combination of genetic, pharmacological, and enzymatic treatment strategies ruled out heparan sulfates and glycoRNA as contributors, although Siglec-8 did exhibit some binding to glycolipids. Genetic disruption of O-mannose glycans within CHST1-expressing cells had a small but significant impact on Siglec-3 and -8 binding, demonstrating that this class of glycans can present sulfated Siglec ligands. We also investigated the ability of sulfated cis ligands to mask Siglec-3 and Siglec-7. For Siglec-7, cis ligands were again found to be mucin-type O-glycans. While N-glycans were the major sulfated trans ligands for Siglec-3, disruption of complex mucin-type O-glycans had the largest impact on Siglec-3 masking. Overall, this study enhances our knowledge of the types of sulfated glycans that can serve as Siglec ligands.","PeriodicalId":11,"journal":{"name":"ACS Chemical Biology","volume":" ","pages":""},"PeriodicalIF":3.5,"publicationDate":"2025-01-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142995974","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Development of an In Situ G Protein-Coupled Receptor Fragment Molecule Screening Approach with High-Resolution Magic Angle Spinning Nuclear Magnetic Resonance. 高分辨率魔角旋转核磁共振原位G蛋白偶联受体片段分子筛选方法的建立。

IF 3.5 2区生物学

ACS Chemical Biology Pub Date : 2025-01-21 DOI: 10.1021/acschembio.4c00686

Enzo Petracco, Guillaume Ferré, Ivo Kabelka, Flavio Ballante, Jens Carlsson, Emma Mulry, Arka P Ray, James Collins, Florent Allais, Matthew T Eddy

{"title":"Development of an In Situ G Protein-Coupled Receptor Fragment Molecule Screening Approach with High-Resolution Magic Angle Spinning Nuclear Magnetic Resonance.","authors":"Enzo Petracco, Guillaume Ferré, Ivo Kabelka, Flavio Ballante, Jens Carlsson, Emma Mulry, Arka P Ray, James Collins, Florent Allais, Matthew T Eddy","doi":"10.1021/acschembio.4c00686","DOIUrl":"https://doi.org/10.1021/acschembio.4c00686","url":null,"abstract":"Small molecules are essential for investigating the pharmacology of membrane proteins and remain the most common approach for therapeutically targeting them. However, most experimental small molecule screening methods require ligands containing radiolabels or fluorescent labels and often involve isolating proteins from their cellular environment. Additionally, most conventional screening methods are suited for identifying compounds with moderate to higher affinities (KD < 1 μM) and are less effective at detecting lower affinity compounds, such as weakly binding molecular fragments. To address these limitations, we demonstrated a proof-of-concept application of high-resolution magic angle spinning nuclear magnetic resonance (HRMAS NMR) spectroscopy with small molecules that bind the human A2A adenosine receptor (A2AAR), a class A G protein-coupled receptor. Our approach leverages a streamlined workflow to prepare NMR samples with only milligrams of unpurified cell membranes containing ∼1 μM of A2AAR. Utilizing saturation transfer difference NMR, we identified bound small molecules from spectra recorded within minutes and further derived information on ligand binding poses without the need for detailed structure determination. After establishing optimal criteria for which the HRMAS approach is most sensitive, we leveraged our HRMAS approach to identify and characterize molecular fragments not previously known to be ligands of A2AAR. In molecular docking and simulations, we observed novel binding poses for these fragments, which revealed the potential to grow them into more complex ligands and confirmed HRMAS NMR as a valuable tool for lead compound identification in the context of fragment-based drug discovery.","PeriodicalId":11,"journal":{"name":"ACS Chemical Biology","volume":" ","pages":""},"PeriodicalIF":3.5,"publicationDate":"2025-01-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142995960","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Breaking the Myth of Enzymatic Azoreduction. 打破酶促偶氮还原的神话。

IF 3.5 2区生物学

ACS Chemical Biology Pub Date : 2025-01-17 Epub Date: 2024-12-21 DOI: 10.1021/acschembio.4c00779

Yu-Ju Peng, Bing Xu, Steven E Rokita

{"title":"Breaking the Myth of Enzymatic Azoreduction.","authors":"Yu-Ju Peng, Bing Xu, Steven E Rokita","doi":"10.1021/acschembio.4c00779","DOIUrl":"10.1021/acschembio.4c00779","url":null,"abstract":"Flavin-dependent azoreductases have been applied to a wide range of tasks from decolorizing numerous azo dyes to releasing azo-conjugated prodrugs. A general narrative reiterated in much of the literature suggests that this enzyme promotes sequential reduction of both the azo-containing substrate and its corresponding hydrazo product to release the aryl amine components while consuming two equivalents of NAD(P)H. Indeed, such aryl amines can be formed by incubation of certain azo compounds with azoreductases, but the nature of the substrates capable of this apparent azo bond lysis remained unknown. We have now prepared a set of azobenzene derivatives and characterized their turnover and products after treatment with azoreductase from Escherichia coli to discover the structural basis regulating aryl amine formation. Without resonance donation by aryl substituents, reduction ceases at the hydrazo product. This indicates that azoreductases do not act on the hydrazo bond. Instead, aryl amine formation depends on a spontaneous hydrazo bond lysis that is promoted by resonance stabilization and subsequent reduction of the quinone-like intermediate by azoreductase. Experimental and computational approaches confirm the substituent dependence of this process. With knowledge of this requirement, full release of aryl amines from azo-conjugates can now be designed and applied with confidence.","PeriodicalId":11,"journal":{"name":"ACS Chemical Biology","volume":" ","pages":"229-237"},"PeriodicalIF":3.5,"publicationDate":"2025-01-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142870570","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Functional Characterization of Pathway Inhibitors for the Ubiquitin-Proteasome System (UPS) as Tool Compounds for CRBN and VHL-Mediated Targeted Protein Degradation. 泛素-蛋白酶体系统（UPS）作为CRBN和vhl介导的靶向蛋白质降解工具化合物的途径抑制剂的功能表征。

IF 3.5 2区生物学

ACS Chemical Biology Pub Date : 2025-01-17 Epub Date: 2025-01-03 DOI: 10.1021/acschembio.4c00450

Martin P Schwalm, Amelie Menge, Lewis Elson, Francesco A Greco, Matthew B Robers, Susanne Müller, Stefan Knapp

{"title":"Functional Characterization of Pathway Inhibitors for the Ubiquitin-Proteasome System (UPS) as Tool Compounds for CRBN and VHL-Mediated Targeted Protein Degradation.","authors":"Martin P Schwalm, Amelie Menge, Lewis Elson, Francesco A Greco, Matthew B Robers, Susanne Müller, Stefan Knapp","doi":"10.1021/acschembio.4c00450","DOIUrl":"10.1021/acschembio.4c00450","url":null,"abstract":"Small molecule degraders such as PROteolysis TArgeting Chimeras (PROTACs) and molecular glues are new modalities for drug development and important tools for target validation. When appropriately optimized, both modalities lead to proteasomal degradation of the protein of interest (POI). Due to the complexity of the induced multistep degradation process, controls for degrader evaluation are critical and commonly used in the literature. However, comparative studies and evaluations of cellular potencies of these control compounds have not been published so far. Here, we investigated a diverse set of ubiquitin pathway inhibitors and evaluated their potency and utility within the CRBN and VHL-mediated degradation pathway. We used the HiBiT system to measure the level of target rescue after treatment with the control compounds. In addition, the cell health was assessed using a multiplexed high-content assay. These assays allowed us to determine nontoxic effective concentrations for control experiments and to perform rescue experiments in the absence of cellular toxicity.","PeriodicalId":11,"journal":{"name":"ACS Chemical Biology","volume":" ","pages":"94-104"},"PeriodicalIF":3.5,"publicationDate":"2025-01-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11745162/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142925797","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","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 Relationship between Substrate Structure and Selectivity of Ketoreduction in Multimodular Polyketide Synthases: A Comparative Study of A-Type Ketoreductases from Late Modules Using Complex Precursor Analogues. 多模聚酮合成酶中底物结构与酮还原选择性的关系：采用复杂前体类似物的后期模A型酮还原酶的比较研究。

IF 3.5 2区生物学

ACS Chemical Biology Pub Date : 2025-01-17 Epub Date: 2025-01-08 DOI: 10.1021/acschembio.4c00669

Lisa N K T Nguyen, Sebastian Derra, Frank Hahn

{"title":"The Relationship between Substrate Structure and Selectivity of Ketoreduction in Multimodular Polyketide Synthases: A Comparative Study of A-Type Ketoreductases from Late Modules Using Complex Precursor Analogues.","authors":"Lisa N K T Nguyen, Sebastian Derra, Frank Hahn","doi":"10.1021/acschembio.4c00669","DOIUrl":"10.1021/acschembio.4c00669","url":null,"abstract":"Ketoreductases (KRs) are domains in the reductive loops of type I polyketide synthases (PKSs) and are responsible for the majority of stereocenters in reduced polyketides. Although the highly stereoselective reduction of ACP-bound β-ketothioester intermediates by KRs is crucial for the overall functioning of PKSs, the substrate-dependent stereoselectivity of KRs is a factor that is not yet fully understood, especially for KR domains in late PKS modules that act on biosynthetic precursors with complex polyketidic moieties. We present studies on the three KR domains FosKR7, PlmKR6, and EryKR6 from the biosynthetic pathways of fostriecin, phoslactomycin, and erythromycin by in vitro assays using close surrogates of the octaketidic FosKR7 biosynthetic precursor, complex derivatives and a diketide in the form of their biomimetic N-acetylcysteamine thioesters. Supported by molecular modeling, specific interactions of the studied KR domains with the extended polyketide moieties of their natural precursors were identified and correlated to the differences in stereoselectivity observed in the in vitro assays. These results reinforce the importance of the substrate-dependent stereoselectivity of KR domains in PKSs and suggest more detailed experimental and structural studies with isolated KRs and full PKS modules that could ultimately lead to improved results in PKS engineering.","PeriodicalId":11,"journal":{"name":"ACS Chemical Biology","volume":" ","pages":"186-196"},"PeriodicalIF":3.5,"publicationDate":"2025-01-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142941420","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Focal Adhesion Regulation as a Strategy against Kidney Fibrosis. 局灶黏附调节作为抗肾纤维化的策略。

IF 3.5 2区生物学

ACS Chemical Biology Pub Date : 2025-01-17 DOI: 10.1021/acschembio.4c00776

Jiwen Geng, Kaikai Zheng, Peng Wang, Baihai Su, Qiang Wei, Xiaojing Liu

引用次数: 0

The Emergence of Oligonucleotide Building Blocks in the Multispecific Proximity-Inducing Drug Toolbox of Destruction. 多特异性接近诱导药物破坏工具箱中寡核苷酸构建块的出现。

IF 3.5 2区生物学

ACS Chemical Biology Pub Date : 2025-01-17 Epub Date: 2024-12-20 DOI: 10.1021/acschembio.4c00311

Kevin Xiao Tong Zhou, Katherine E Bujold

{"title":"The Emergence of Oligonucleotide Building Blocks in the Multispecific Proximity-Inducing Drug Toolbox of Destruction.","authors":"Kevin Xiao Tong Zhou, Katherine E Bujold","doi":"10.1021/acschembio.4c00311","DOIUrl":"10.1021/acschembio.4c00311","url":null,"abstract":"Oligonucleotides are a rapidly emerging class of therapeutics. Their most well-known examples are informational drugs that modify gene expression by binding mRNA. Despite inducing proximity between biological machinery and mRNA when applied to modulating gene expression, oligonucleotides are not typically labeled as \"proximity-inducing\" in literature. Yet, they have recently been explored as building blocks for multispecific proximity-inducing drugs (MPIDs). MPIDs are unique because they can direct endogenous biological machinery to destroy targeted molecules and cells, in contrast to traditional drugs that inhibit only their functions. The unique mechanism of action of MPIDs has enabled the targeting of previously \"undruggable\" molecular entities that cannot be effectively inhibited. However, the development of MPIDs must ensure that these molecules will selectively direct a potent, destruction-based mechanism of action toward intended targets over healthy tissues to avoid causing life-threatening toxicities. Oligonucleotides have emerged as promising building blocks for the design of MPIDs because they are sequence-controlled molecules that can be rationally designed to program multispecific binding interactions. In this Review, we examine the emergence of oligonucleotide-containing MPIDs in the proximity induction space, which has been dominated by antibody and small molecule MPID modalities. Moreover, examples of oligonucleotides developed as MPID candidates in immunotherapy and protein degradation are discussed to demonstrate the utility of oligonucleotides in expanding the scope and selectivity of the MPID toolbox. Finally, we discuss the utility of programming \"AND\" gates into oligonucleotide scaffolds to encode conditional responses that have the potential to be incorporated into MPIDs, which can further enhance their selectivity, thus increasing the scope of this drug category.","PeriodicalId":11,"journal":{"name":"ACS Chemical Biology","volume":" ","pages":"3-18"},"PeriodicalIF":3.5,"publicationDate":"2025-01-17","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142862500","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":2,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Novel Quinazoline Derivatives Inhibit Splicing of Fungal Group II Introns. 新型喹唑啉衍生物抑制真菌II族内含子剪接。

IF 3.5 2区生物学

ACS Chemical Biology Pub Date : 2025-01-17 DOI: 10.1021/acschembio.4c00631

Olga Fedorova, Michelle Luo, G Erik Jagdmann, Michael C Van Zandt, Luke Sisto, Anna Marie Pyle

引用次数: 0