ACS Chemical Biology最新文献

A C-Degron Structure-Based Approach for the Development of Ligands Targeting the E3 Ligase TRIM7. 基于 C-egron 结构的配体靶向 E3 配体 TRIM7 的开发方法。

IF 3.5 2区生物学

ACS Chemical Biology Pub Date : 2024-06-27 DOI: 10.1021/acschembio.4c00301

Christian J Muñoz Sosa, Christopher Lenz, Anton Hamann, Frederic Farges, Johannes Dopfer, Andreas Krämer, Veronika Cherkashyna, Andrey Tarnovskiy, Yurii S Moroz, Ewgenij Proschak, Václav Němec, Susanne Müller, Krishna Saxena, Stefan Knapp

{"title":"A C-Degron Structure-Based Approach for the Development of Ligands Targeting the E3 Ligase TRIM7.","authors":"Christian J Muñoz Sosa, Christopher Lenz, Anton Hamann, Frederic Farges, Johannes Dopfer, Andreas Krämer, Veronika Cherkashyna, Andrey Tarnovskiy, Yurii S Moroz, Ewgenij Proschak, Václav Němec, Susanne Müller, Krishna Saxena, Stefan Knapp","doi":"10.1021/acschembio.4c00301","DOIUrl":"https://doi.org/10.1021/acschembio.4c00301","url":null,"abstract":"TRIM7 is a ubiquitin E3 ligase with key regulatory functions, mediating viral infection, tumor biology, innate immunity, and cellular processes, such as autophagy and ferroptosis. It contains a PRYSPRY domain that specifically recognizes degron sequences containing C-terminal glutamine. Ligands that bind to the TRIM7 PRYSPRY domain may have applications in the treatment of viral infections, as modulators of inflammation, and in the design of a new class of PROTACs (PROteolysis TArgeting Chimeras) that mediate the selective degradation of therapeutically relevant proteins (POIs). Here, we developed an assay toolbox for the comprehensive evaluation of TRIM7 ligands. Using TRIM7 degron sequences together with a structure-based design, we developed the first series of peptidomimetic ligands with low micromolar affinity. The terminal carboxylate moiety was required for ligand activity but prevented cell penetration. A prodrug strategy using an ethyl ester resulted in enhanced permeability, which was evaluated using confocal imaging.","PeriodicalId":11,"journal":{"name":"ACS Chemical Biology","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-06-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141453610","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

Establishing a Cell-Free Glycoprotein Synthesis System for Enzymatic N-GlcNAcylation.

IF 3.5 2区生物学

ACS Chemical Biology Pub Date : 2024-06-27 DOI: 10.1021/acschembio.4c00228

Madison A DeWinter, Derek A Wong, Regina Fernandez, Weston Kightlinger, Ariel Helms Thames, Matthew P DeLisa, Michael C Jewett

{"title":"Establishing a Cell-Free Glycoprotein Synthesis System for Enzymatic N-GlcNAcylation.","authors":"Madison A DeWinter, Derek A Wong, Regina Fernandez, Weston Kightlinger, Ariel Helms Thames, Matthew P DeLisa, Michael C Jewett","doi":"10.1021/acschembio.4c00228","DOIUrl":"https://doi.org/10.1021/acschembio.4c00228","url":null,"abstract":"N-linked glycosylation plays a key role in the efficacy of many therapeutic proteins. One limitation to the bacterial glycoengineering of human N-linked glycans is the difficulty of installing a single N-acetylglucosamine (GlcNAc), the reducing end sugar of many human-type glycans, onto asparagine in a single step (N-GlcNAcylation). Here, we develop an in vitro method for N-GlcNAcylating proteins using the oligosaccharyltransferase PglB from Campylobacter jejuni. We use cell-free protein synthesis (CFPS) to test promiscuous PglB variants previously reported in the literature for the ability to produce N-GlcNAc and successfully determine that PglB with an N311V mutation (PglBN311V) exhibits increased GlcNAc transferase activity relative to the wild-type enzyme. We then improve the transfer efficiency by producing CFPS extracts enriched with PglBN311V and further optimize the reaction conditions, achieving a 98.6 ± 0.5% glycosylation efficiency. We anticipate this method will expand the glycoengineering toolbox for therapeutic research and biomanufacturing.","PeriodicalId":11,"journal":{"name":"ACS Chemical Biology","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-06-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141453611","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

N-Acyl-N-Alkyl Sulfonamide Probes for Ligand-Directed Covalent Labeling of GPCRs: The Adenosine A_2B Receptor as Case Study. 用于配体引导共价标记 GPCR 的 N-酰基-N-烷基磺酰胺探针：腺苷 A2B 受体案例研究。

IF 3.5 2区生物学

ACS Chemical Biology Pub Date : 2024-06-26 DOI: 10.1021/acschembio.4c00210

Bert L H Beerkens, Vasiliki Andrianopoulou, Xuesong Wang, Rongfang Liu, Gerard J P van Westen, Willem Jespers, Adriaan P IJzerman, Laura H Heitman, Daan van der Es

{"title":"N-Acyl-N-Alkyl Sulfonamide Probes for Ligand-Directed Covalent Labeling of GPCRs: The Adenosine A2B Receptor as Case Study.","authors":"Bert L H Beerkens, Vasiliki Andrianopoulou, Xuesong Wang, Rongfang Liu, Gerard J P van Westen, Willem Jespers, Adriaan P IJzerman, Laura H Heitman, Daan van der Es","doi":"10.1021/acschembio.4c00210","DOIUrl":"https://doi.org/10.1021/acschembio.4c00210","url":null,"abstract":"Small molecular tool compounds play an essential role in the study of G protein-coupled receptors (GPCRs). However, tool compounds most often occupy the orthosteric binding site, hampering the study of GPCRs upon ligand binding. To overcome this problem, ligand-directed labeling techniques have been developed that leave a reporter group covalently bound to the GPCR, while allowing subsequent orthosteric ligands to bind. In this work, we applied such a labeling strategy to the adenosine A2B receptor (A2BAR). We have synthetically implemented the recently reported N-acyl-N-alkyl sulfonamide (NASA) warhead into a previously developed ligand and show that the binding of the A2BAR is not restricted by NASA incorporation. Furthermore, we have investigated ligand-directed labeling of the A2BAR using SDS-PAGE, flow cytometric, and mass spectrometry techniques. We have found one of the synthesized probes to specifically label the A2BAR, although detection was hindered by nonspecific protein labeling most likely due to the intrinsic reactivity of the NASA warhead. Altogether, this work aids the future development of ligand-directed probes for the detection of GPCRs.","PeriodicalId":11,"journal":{"name":"ACS Chemical Biology","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-06-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141449019","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

De Novo Synthesis and Structural Elucidation of CDR-H3 Loop Mimics. CDR-H3环路模拟物的新合成与结构阐明

IF 3.5 2区生物学

ACS Chemical Biology Pub Date : 2024-06-25 DOI: 10.1021/acschembio.4c00236

Guangkuan Zhao, Alexis D Richaud, R Thomas Williamson, Michael Feig, Stéphane P Roche

{"title":"De Novo Synthesis and Structural Elucidation of CDR-H3 Loop Mimics.","authors":"Guangkuan Zhao, Alexis D Richaud, R Thomas Williamson, Michael Feig, Stéphane P Roche","doi":"10.1021/acschembio.4c00236","DOIUrl":"https://doi.org/10.1021/acschembio.4c00236","url":null,"abstract":"The binding affinity of antibodies to specific antigens stems from a remarkably broad repertoire of hypervariable loops known as complementarity-determining regions (CDRs). While recognizing the pivotal role of the heavy-chain 3 CDRs (CDR-H3s) in maximizing antibody-antigen affinity and specificity, the key structural determinants responsible for their adaptability to diverse loop sequences, lengths, and noncanonical structures are hitherto unknown. To address this question, we achieved a de novo synthesis of bulged CDR-H3 mimics excised from their full antibody context. CD and NMR data revealed that these stable standalone β-hairpin scaffolds are well-folded and retain many of the native bulge CDR-H3 features in water. In particular, the tryptophan residue, highly conserved across CDR-H3 sequences, was found to extend the kinked base of these β-bulges through a combination of stabilizing intramolecular hydrogen bond and CH/π interaction. The structural ensemble consistent with our NMR observations exposed the dynamic nature of residues at the base of the loop, suggesting that β-bulges act as molecular hinges connecting the rigid stem to the more flexible loops of CDR-H3s. We anticipate that this deeper structural understanding of CDR-H3s will lay the foundation to inform the design of antibody drugs broadly and engineer novel CDR-H3 peptide scaffolds as therapeutics.","PeriodicalId":11,"journal":{"name":"ACS Chemical Biology","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-06-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141445442","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

Native Aminoacyl-tRNA Synthetase/tRNA Pair Drives Highly Efficient Noncanonical Amino Acid Incorporation in Escherichia coli. 大肠杆菌中的原生氨基酰-tRNA 合成酶/tRNA 对驱动了高效的非规范氨基酸结合。

IF 3.5 2区生物学

ACS Chemical Biology Pub Date : 2024-06-24 DOI: 10.1021/acschembio.4c00221

Elise D Ficaretta, Soumya Jyoti Singha Roy, Lena Voss, Abhishek Chatterjee

{"title":"Native Aminoacyl-tRNA Synthetase/tRNA Pair Drives Highly Efficient Noncanonical Amino Acid Incorporation in Escherichia coli.","authors":"Elise D Ficaretta, Soumya Jyoti Singha Roy, Lena Voss, Abhishek Chatterjee","doi":"10.1021/acschembio.4c00221","DOIUrl":"https://doi.org/10.1021/acschembio.4c00221","url":null,"abstract":"Site-specific noncanonical amino acid (ncAA) mutagenesis in living cells has traditionally relied on heterologous, nonsense-suppressing aminoacyl-tRNA synthetase (aaRS)/tRNA pairs that do not cross-react with their endogenous counterparts. Such heterologous pairs often perform suboptimally in a foreign host cell since they were not evolutionarily optimized to function in the foreign environment. This suboptimal performance restricts the number of ncAAs that can be simultaneously incorporated into a protein. Here, we show that the use of an endogenous aaRS/tRNA pair to drive ncAA incorporation can offer a potential solution to this limitation. To this end, we developed an engineered Escherichia coli strain (ATMY-C321), wherein the endogenous tyrosyl-tRNA synthetase (TyrRS)/tRNA pair has been functionally replaced with an archaeal counterpart, and the release factor 1 has been removed to eliminate competing termination at the UAG nonsense codons. The endogenous TyrRS/tRNACUATyr pair exhibits remarkably efficient nonsense suppression in the resulting cell, relative to established orthogonal ncAA-incorporation systems in E. coli, allowing the incorporation of an ncAA at up to 10 contiguous sites in a reporter protein. Our work highlights the limitations of orthogonal translation systems using heterologous aaRS/tRNA pairs and offers a potential alternative involving the use of endogenous pairs.","PeriodicalId":11,"journal":{"name":"ACS Chemical Biology","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-06-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141445443","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

A Robust α-l-Fucosidase from Prevotella nigrescens for Glycoengineering Therapeutic Antibodies. 从黑前孢子菌中提取的用于糖工程治疗抗体的强效α-l-岩藻糖苷酶

IF 3.5 2区生物学

ACS Chemical Biology Pub Date : 2024-06-24 DOI: 10.1021/acschembio.4c00196

Mu-Rong Kao, Tzu-Hsuan Ma, Hsiang-Yu Chou, Shu-Chieh Chang, Lin-Chen Cheng, Kuo-Shiang Liao, Jiun-Jie Shie, Philip J Harris, Chi-Huey Wong, Yves S Y Hsieh

{"title":"A Robust α-l-Fucosidase from Prevotella nigrescens for Glycoengineering Therapeutic Antibodies.","authors":"Mu-Rong Kao, Tzu-Hsuan Ma, Hsiang-Yu Chou, Shu-Chieh Chang, Lin-Chen Cheng, Kuo-Shiang Liao, Jiun-Jie Shie, Philip J Harris, Chi-Huey Wong, Yves S Y Hsieh","doi":"10.1021/acschembio.4c00196","DOIUrl":"https://doi.org/10.1021/acschembio.4c00196","url":null,"abstract":"Eliminating the core fucose from the N-glycans of the Fc antibody segment by pathway engineering or enzymatic methods has been shown to enhance the potency of therapeutic antibodies, especially in the context of antibody-dependent cytotoxicity (ADCC). However, there is a significant challenge due to the limited defucosylation efficiency of commercially available α-l-fucosidases. In this study, we report a unique α-l-fucosidase (PnfucA) from the bacterium Prevotella nigrescens that has a low sequence identity compared with all other known α-l-fucosidases and is highly reactive toward a core disaccharide substrate with fucose α(1,3)-, α (1,4)-and α(1,6)-linked to GlcNAc, and is less reactive toward the Fuc-α(1,2)-Gal on the terminal trisaccharide of the oligosaccharide Globo H (Bb3). The kinetic properties of the enzyme, such as its Km and kcat, were determined and the optimized expression of PnfucA gave a yield exceeding 30 mg/L. The recombinant enzyme retained its full activity even after being incubated for 6 h at 37 °C. Moreover, it retained 92 and 87% of its activity after freezing and freeze-drying treatments, respectively, for over 28 days. In a representative glycoengineering of adalimumab (Humira), PnfucA showed remarkable hydrolytic efficiency in cleaving the α(1,6)-linked core fucose from FucGlcNAc on the antibody with a quantitative yield. This enabled the seamless incorporation of biantennary sialylglycans by Endo-S2 D184 M in a one-pot fashion to yield adalimumab in a homogeneous afucosylated glycoform with an improved binding affinity toward Fcγ receptor IIIa.","PeriodicalId":11,"journal":{"name":"ACS Chemical Biology","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-06-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141441697","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

Phosphinate Esters as Novel Warheads for Quenched Activity-Based Probes Targeting Serine Proteases. 磷酸盐酯作为新型弹头，用于淬火活性探针靶向丝氨酸蛋白酶。

IF 3.5 2区生物学

ACS Chemical Biology Pub Date : 2024-06-24 DOI: 10.1021/acschembio.3c00203

Jan Pascal Kahler, Shanping Ji, Femke Speelman-Rooms, Roeland Vanhoutte, Steven H L Verhelst

引用次数: 0

Characterization, Structure, and Inhibition of the Human Succinyl-CoA:glutarate-CoA Transferase, a Putative Genetic Modifier of Glutaric Aciduria Type 1. 人类琥珀酰-CoA:戊二酸-CoA 转化酶的特性、结构和抑制作用，它是戊二酸尿症 1 型的推定遗传修饰因子。

IF 3.5 2区生物学

ACS Chemical Biology Pub Date : 2024-06-24 DOI: 10.1021/acschembio.4c00204

Ruoxi Wu, Susmita Khamrui, Tetyana Dodatko, João Leandro, Amanda Sabovic, Sara Violante, Justin R Cross, Eric Marsan, Kunal Kumar, Robert J DeVita, Michael B Lazarus, Sander M Houten

{"title":"Characterization, Structure, and Inhibition of the Human Succinyl-CoA:glutarate-CoA Transferase, a Putative Genetic Modifier of Glutaric Aciduria Type 1.","authors":"Ruoxi Wu, Susmita Khamrui, Tetyana Dodatko, João Leandro, Amanda Sabovic, Sara Violante, Justin R Cross, Eric Marsan, Kunal Kumar, Robert J DeVita, Michael B Lazarus, Sander M Houten","doi":"10.1021/acschembio.4c00204","DOIUrl":"10.1021/acschembio.4c00204","url":null,"abstract":"Glutaric Aciduria Type 1 (GA1) is a serious inborn error of metabolism with no pharmacological treatments. A novel strategy to treat this disease is to divert the toxic biochemical intermediates to less toxic or nontoxic metabolites. Here, we report a putative novel target, succinyl-CoA:glutarate-CoA transferase (SUGCT), which we hypothesize suppresses the GA1 metabolic phenotype through decreasing glutaryl-CoA and the derived 3-hydroxyglutaric acid. SUGCT is a type III CoA transferase that uses succinyl-CoA and glutaric acid as substrates. We report the structure of SUGCT, develop enzyme- and cell-based assays, and identify valsartan and losartan carboxylic acid as inhibitors of the enzyme in a high-throughput screen of FDA-approved compounds. The cocrystal structure of SUGCT with losartan carboxylic acid revealed a novel pocket in the active site and further validated the high-throughput screening approach. These results may form the basis for the future development of new pharmacological intervention to treat GA1.","PeriodicalId":11,"journal":{"name":"ACS Chemical Biology","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-06-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141445441","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

Prokaryotic RNA N1-Methyladenosine Erasers Maintain tRNA m1A Modification Levels in Streptomyces venezuelae. 原核 RNA N1-甲基腺苷清除剂可维持委内瑞拉链霉菌中 tRNA m1A 的修饰水平。

IF 3.5 2区生物学

ACS Chemical Biology Pub Date : 2024-06-24 DOI: 10.1021/acschembio.4c00278

Marcus Foo, Luke R Frietze, Behnam Enghiad, Yujie Yuan, Christopher D Katanski, Huimin Zhao, Tao Pan

{"title":"Prokaryotic RNA N1-Methyladenosine Erasers Maintain tRNA m1A Modification Levels in Streptomyces venezuelae.","authors":"Marcus Foo, Luke R Frietze, Behnam Enghiad, Yujie Yuan, Christopher D Katanski, Huimin Zhao, Tao Pan","doi":"10.1021/acschembio.4c00278","DOIUrl":"https://doi.org/10.1021/acschembio.4c00278","url":null,"abstract":"tRNA modifications help maintain tRNA structure and facilitate translation and stress response. Found in all three kingdoms of life, m1A tRNA modification occurs in the T loop of many tRNAs, stabilizes tertiary tRNA structure, and impacts translation. M1A in the T loop is reversible by three mammalian demethylase enzymes, which bypasses the need of turning over the tRNA molecule to adjust its m1A levels in cells. However, no prokaryotic tRNA demethylase enzyme has been identified that acts on endogenous RNA modifications. Using Streptomyces venezuelae as a model organism, we confirmed the presence and quantitative m1A tRNA signatures using mass spectrometry and high-throughput tRNA sequencing. We identified two RNA demethylases that can remove m1A in tRNA and validated the activity of a previously annotated tRNA m1A writer. Using single-gene knockouts of these erasers and the m1A writer, we found dynamic changes of m1A levels in many tRNAs under stress conditions. Phenotypic characterization highlighted changes in their growth and altered antibiotic production. Our identification of the first prokaryotic tRNA demethylase enzyme paves the way for investigating new mechanisms of translational regulation in bacteria.","PeriodicalId":11,"journal":{"name":"ACS Chemical Biology","volume":null,"pages":null},"PeriodicalIF":3.5,"publicationDate":"2024-06-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"141441698","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

Fluorometric Analysis of Carrier-Protein-Dependent Biosynthesis through a Conformationally Sensitive Solvatochromic Pantetheinamide Probe. 通过构象敏感的溶解变色泛硫乙酰胺探针，对载体蛋白依赖性生物合成进行荧光分析。

IF 3.5 2区生物学

ACS Chemical Biology Pub Date : 2024-06-23 DOI: 10.1021/acschembio.4c00169

Matthew G Miyada, Yuran Choi, Ramunas Stepanauskas, Tanja Woyke, James J La Clair, Michael D Burkart

引用次数: 0