Biochemistry Biochemistry最新文献_第3页

Coupling Subunit-Specific States to Allosteric Regulation in Homodimeric Cyclooxygenase-2.

IF 2.9 3区生物学

Biochemistry Biochemistry Pub Date : 2025-02-28 DOI: 10.1021/acs.biochem.4c00821

Liang Dong, Michael G Malkowski

{"title":"Coupling Subunit-Specific States to Allosteric Regulation in Homodimeric Cyclooxygenase-2.","authors":"Liang Dong, Michael G Malkowski","doi":"10.1021/acs.biochem.4c00821","DOIUrl":"https://doi.org/10.1021/acs.biochem.4c00821","url":null,"abstract":"The homodimeric cyclooxygenase enzymes (COX-1 and COX-2) oxygenate arachidonic acid (AA) to generate prostaglandins. COX-2 behaves as a conformational heterodimer in solution comprised of allosteric (Eallo) and catalytic (Ecat) subunits that function cooperatively. We previously utilized 19F-nuclear magnetic resonance spectroscopy (19F-NMR) to show that the cyclooxygenase active site entrances in a COX-2 homodimer construct exhibited composite tightened and relaxed states that are dependent upon the type of ligand bound. A third state, hypothesized to represent the alteration of a loop comprised of residues 120-129, was also detected in the presence of ligands that allosterically potentiate activity. We report here studies that couple the use of 19F-NMR with COX-2 heterodimer constructs to characterize states arising in the individual subunits. Glycine and proline substitutions at Ser-121 were introduced to examine how these mutations alter the 120-129 loop. In the presence of AA, the subunits exhibited asymmetry, with tightened and relaxed states observed in Eallo and Ecat, respectively. Allosteric ligand binding resulted in a shift to equivalent symmetrical states, with tightened states observed in the presence of the allosteric inhibitor flurbiprofen and relaxed states observed in the presence of the allosteric potentiator palmitic acid. The S121P substitution results in a shift to equivalent relaxed states, as well as an alteration of the 120-129 loop in the absence of bound ligand. We put forth a model linking the observed differential states arising from allosteric ligand binding with structural transitions across the dimer interface that govern the regulation of cyclooxygenase activity.","PeriodicalId":28,"journal":{"name":"Biochemistry Biochemistry","volume":" ","pages":""},"PeriodicalIF":2.9,"publicationDate":"2025-02-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143530872","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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

Decoding Hairpin Structure Stability in Lin28-Mediated Repression.

IF 2.9 3区生物学

Biochemistry Biochemistry Pub Date : 2025-02-28 DOI: 10.1021/acs.biochem.4c00675

Qiang Zhu, Limu Hu, Chang Cui, Min Zang, Hao Dong, Jing Ma

{"title":"Decoding Hairpin Structure Stability in Lin28-Mediated Repression.","authors":"Qiang Zhu, Limu Hu, Chang Cui, Min Zang, Hao Dong, Jing Ma","doi":"10.1021/acs.biochem.4c00675","DOIUrl":"https://doi.org/10.1021/acs.biochem.4c00675","url":null,"abstract":"The Lin28 protein is well known for its role in inhibiting the biogenesis of microRNAs (miRNAs) that belong to the let-7 family. The Lin28 and let-7 axes are associated with several types of cancers. It is imperative to understand the underlying mechanism to treat these cancers in a more efficient way. In this study, we employed all-atom molecular dynamics simulation as a research tool to investigate the interaction formed between Lin28 and the precursor element of let-7d, one of the 12 members of the let-7 family. By constructing systems of an intact sequence length of preE-let-7d, our simulations suggest that both the loop region of the hairpin structure and the GGAG sequence can form stable interactions with the cold shock domain (CSD) and zinc knuckle domain (ZKD) regions of the protein, respectively. The system, by deleting the nucleotides GGAG at the 3' terminal, indicates that the loop region is more responsible for its ability in bypassing the binding and repression of Lin28. Additionally, using let-7c-2, which can bypass Lin28 regulation, as a template, we constructed systems with mutated loop region sequences in miRNAs and tested their stabilities. Our simulation results coincide well with experimental observations. Based on both simulation results and statistical analysis from two databases, we hypothesized that two factors, namely, the interaction between terminal nucleotides and the ring tension originating from the middle nucleotides, can significantly influence their stabilities. Systems combining strong and weak terminal interactions with large and small ring tensions were recruited to validate our hypothesis. Our findings offer a new perspective and shed light on strategies for designing sequences to regulate the interactions formed between proteins and hairpin structures.","PeriodicalId":28,"journal":{"name":"Biochemistry Biochemistry","volume":" ","pages":""},"PeriodicalIF":2.9,"publicationDate":"2025-02-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143527804","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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

Integrated Study of Fluorescence Enhancement in the Y176H Variant of Cyanobacterial Phytochrome Cph1. 蓝藻植物色素 Cph1 Y176H 变体的荧光增强综合研究

IF 2.9 3区生物学

Biochemistry Biochemistry Pub Date : 2025-02-27 DOI: 10.1021/acs.biochem.4c00687

Soshichiro Nagano, Chen Song, Valentin Rohr, Megan J Mackintosh, Oanh Tu Hoang, Anastasia Kraskov, Yang Yang, Jon Hughes, Karsten Heyne, Maria-Andrea Mroginski, Igor Schapiro, Peter Hildebrandt

{"title":"Integrated Study of Fluorescence Enhancement in the Y176H Variant of Cyanobacterial Phytochrome Cph1.","authors":"Soshichiro Nagano, Chen Song, Valentin Rohr, Megan J Mackintosh, Oanh Tu Hoang, Anastasia Kraskov, Yang Yang, Jon Hughes, Karsten Heyne, Maria-Andrea Mroginski, Igor Schapiro, Peter Hildebrandt","doi":"10.1021/acs.biochem.4c00687","DOIUrl":"https://doi.org/10.1021/acs.biochem.4c00687","url":null,"abstract":"Phytochromes are red-light-sensitive biliprotein photoreceptors that control a variety of physiological processes in plants, fungi, and bacteria. Lately, greater attention has been paid to these photoreceptors due to their potential as fluorescent probes for deep-tissue microscopy. Such fluorescing phytochromes have been generated by multiple amino acid substitutions in weakly fluorescent wild-type (WT) proteins. Remarkably, the single substitution of conserved Tyr176 by His in cyanobacterial phytochrome Cph1 increases the fluorescence quantum yield from 2.4 to 14.5%. In this work, we studied this Y176H variant by crystallography, MAS NMR, resonance Raman spectroscopy, and ultrafast absorption spectroscopy complemented by theoretical methods. Two factors were identified to account for the strong fluorescence increase. First, the equilibrium between the photoactive and fluorescent substates of WT Cph1 was shown to shift entirely to the fluorescent substate in Y176H. Second, structural flexibility of the chromophore is drastically reduced and the photoisomerization barrier is raised, thereby increasing the excited-state lifetime. The most striking finding, however, is that Y176H includes the structural properties of both the dark-adapted Pr and the light-activated Pfr state. While the chromophore adopts the Pr-typical ZZZssa configuration, the tongue segment of the protein adopts a Pfr-typical α-helical structure. This implies that Tyr176 plays a key role in coupling chromophore photoisomerization to the sheet-to-helix transition of the tongue and the final Pfr structure. This conclusion extends to plant phytochromes, where the homologous substitution causes light-independent signaling activity akin to that of Pfr.","PeriodicalId":28,"journal":{"name":"Biochemistry Biochemistry","volume":" ","pages":""},"PeriodicalIF":2.9,"publicationDate":"2025-02-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143522198","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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

Biased GPCR Signaling: Possible Mechanisms and Therapeutic Applications. 偏性 GPCR 信号：可能的机制和治疗应用。

IF 2.9 3区生物学

Biochemistry Biochemistry Pub Date : 2025-02-27 DOI: 10.1021/acs.biochem.4c00827

Luyu Fan, Sheng Wang

{"title":"Biased GPCR Signaling: Possible Mechanisms and Therapeutic Applications.","authors":"Luyu Fan, Sheng Wang","doi":"10.1021/acs.biochem.4c00827","DOIUrl":"https://doi.org/10.1021/acs.biochem.4c00827","url":null,"abstract":"Biased signaling refers to the phenomenon where a ligand selectively activates specific downstream pathways of G protein-coupled receptors (GPCRs), such as the G protein-mediated pathway or the β-arrestin-mediated pathway. This mechanism can be influenced by receptor bias, ligand bias, system bias and spatial bias, all of which are shaped by the receptor's conformational distinctions and kinetics. Since GPCRs are the largest class of drug targets, signaling bias garnered significant attention for its potential to enhance therapeutic efficacy while minimizing side effects. Despite intensive investigation, a major challenge lies in translating in vitro ligand efficacy into in vivo biological responses due to the dynamic and multifaceted nature of the in vivo environment. This review delves into the current understanding of GPCR-biased signaling, examining the role of structural bias at the molecular level, the impact of kinetic context on system and observational bias, and the challenges of applying these insights in drug development. It further explores future directions for advancing biased signaling applications, offering valuable perspectives on how to bridge the gap between in vitro studies and in vivo therapeutic design, ultimately accelerating the development of viable, biased therapeutics.","PeriodicalId":28,"journal":{"name":"Biochemistry Biochemistry","volume":" ","pages":""},"PeriodicalIF":2.9,"publicationDate":"2025-02-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143522197","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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

Targeting Bacterial RNA Polymerase: Harnessing Simulations and Machine Learning to Design Inhibitors for Drug-Resistant Pathogens.

IF 2.9 3区生物学

Biochemistry Biochemistry Pub Date : 2025-02-27 DOI: 10.1021/acs.biochem.4c00751

Eshani C Goonetilleke, Xuhui Huang

{"title":"Targeting Bacterial RNA Polymerase: Harnessing Simulations and Machine Learning to Design Inhibitors for Drug-Resistant Pathogens.","authors":"Eshani C Goonetilleke, Xuhui Huang","doi":"10.1021/acs.biochem.4c00751","DOIUrl":"https://doi.org/10.1021/acs.biochem.4c00751","url":null,"abstract":"The increase in antimicrobial resistance presents a major challenge in treating bacterial infections, underscoring the need for innovative drug discovery approaches and novel inhibitors. Bacterial RNA polymerase (RNAP) has emerged as a crucial target for antibiotic development due to its essential role in transcription. RNAP is a molecular motor, and its function relies heavily on the dynamic shifts between multiple conformational states. While biochemical and structural experimental methods offer crucial insights into static RNAP-drug interactions, they fall short in capturing the dynamics at a molecular level. By integrating experimental data with advanced computational techniques like Markov State Models (MSMs), Generalized Master Equation (GME) Models and other machine-learning models constructed from molecular dynamics (MD) simulations, researchers can elucidate novel cryptic pockets that open transiently for antibiotic compounds and gain a more nuanced and comprehensive understanding of RNAP-drug interactions. This integrated approach not only deepens our fundamental knowledge but also enables more targeted and efficient antibiotic design strategies. In this Perspective, we highlight how this synergy between experimental and computational methods has the potential to open new pathways for innovative drug design and combination therapies that may help turn the tide in the ongoing battle against antibiotic-resistant bacteria.","PeriodicalId":28,"journal":{"name":"Biochemistry Biochemistry","volume":" ","pages":""},"PeriodicalIF":2.9,"publicationDate":"2025-02-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143514005","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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

Tumor Microenvironment pH-Sensitive Peptidomimetics for Targeted Anticancer Drug Delivery. 用于靶向递送抗癌药物的肿瘤微环境 pH 值敏感拟肽物

IF 2.9 3区生物学

Biochemistry Biochemistry Pub Date : 2025-02-27 DOI: 10.1021/acs.biochem.4c00657

Biswanath Maity, Hariharan Moorthy, Thimmaiah Govindaraju

{"title":"Tumor Microenvironment pH-Sensitive Peptidomimetics for Targeted Anticancer Drug Delivery.","authors":"Biswanath Maity, Hariharan Moorthy, Thimmaiah Govindaraju","doi":"10.1021/acs.biochem.4c00657","DOIUrl":"https://doi.org/10.1021/acs.biochem.4c00657","url":null,"abstract":"Cell-penetrating peptides (CPPs) are known for their effective intracellular transport of bioactives such as therapeutic proteins, peptides, nucleic acid, and small molecule drugs. However, the excessive cationic charges that promote their membrane permeability result in nonselective delivery and cellular toxicity. In this study, we report a decamer cell-penetrating peptidomimetic, Hkd, designed to selectively deliver anticancer drugs into tumor cells in response to the acidic microenvironment. The pH-sensitive histidine (H) imidazole side chain undergoes protonation in acidic environments, facilitating membrane permeability. The rigid cyclic dipeptide (CDP) core (kd) of Hkd has multiple hydrogen bond donor and acceptor sites, enabling selective interaction-driven cellular uptake. Pharmacokinetic studies revealed the excellent serum stability of Hkd. Cellular uptake studies of Hkd showed improved uptake at a lower pH than physiological pH. Conjugation of Hkd to the anticancer drug camptothecin (Cpt) reduced nonselective drug transport to normal cells while effectively delivering the drug into cancerous cells at the tumor microenvironment pH and retaining the therapeutic potential of the drug. The systematic design of pH-sensitive peptidomimetics offers a viable method to overcome the challenges of stability and selectivity faced by traditional highly cationic CPPs, potentially expanding the application range of this delivery system.","PeriodicalId":28,"journal":{"name":"Biochemistry Biochemistry","volume":" ","pages":""},"PeriodicalIF":2.9,"publicationDate":"2025-02-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143522199","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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 "Knob Switch" Model for the Phosphoregulatory Mechanism of KCC3 at the Carboxy-Terminal Domain.

IF 2.9 3区生物学

Biochemistry Biochemistry Pub Date : 2025-02-26 DOI: 10.1021/acs.biochem.4c00523

Xiaoli Lu, Jing Xue, Qiang Zhou, Jing Huang

{"title":"A \"Knob Switch\" Model for the Phosphoregulatory Mechanism of KCC3 at the Carboxy-Terminal Domain.","authors":"Xiaoli Lu, Jing Xue, Qiang Zhou, Jing Huang","doi":"10.1021/acs.biochem.4c00523","DOIUrl":"https://doi.org/10.1021/acs.biochem.4c00523","url":null,"abstract":"Phosphorylation is a reversible post-translational modification that can modulate protein function. For example, phosphorylation modifications of solute carrier family 12 (SLC12) proteins function as molecular switches that precisely regulate cation-chloride ion transport. Elucidating the phosphoregulatory mechanism of SLC12 at the carboxy-terminal domain (CTD) through structural determination approaches remains challenging due to the domain's disordered and flexible nature. In this study, molecular dynamics (MD) simulations and enhanced sampling techniques were employed to investigate the CTD phosphoregulatory mechanism of SLC12A6 (also known as KCC3). Atomistic MD and metadynamics simulations revealed that the dephosphorylation of residues T940 and T997 stabilizes CTD to a favorable state that \"switches on\" the solvent accessibility of the inward-facing pocket. Meanwhile, phosphorylation induces distinct orientations of the CTD, transitioning the dimer into another favorable state that \"switches off\" the solvent accessibility. The alteration of solvent accessibility in the inward-facing pocket influences the water and ion dynamics. Based on these findings, we propose a \"knob switch\" model to illustrate how CTD phosphorylation regulates ion transport in KCC3.","PeriodicalId":28,"journal":{"name":"Biochemistry Biochemistry","volume":" ","pages":""},"PeriodicalIF":2.9,"publicationDate":"2025-02-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143513999","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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

Phase Separation Clustering of Poly Ubiquitin Cargos on Ternary Mixture Lipid Membranes by Synthetically Cross-Linked Ubiquitin Binder Peptides.

IF 2.9 3区生物学

Biochemistry Biochemistry Pub Date : 2025-02-26 DOI: 10.1021/acs.biochem.4c00483

Soojung Kim, Kamsy K Okafor, Rina Tabuchi, Cedric Briones, Il-Hyung Lee

{"title":"Phase Separation Clustering of Poly Ubiquitin Cargos on Ternary Mixture Lipid Membranes by Synthetically Cross-Linked Ubiquitin Binder Peptides.","authors":"Soojung Kim, Kamsy K Okafor, Rina Tabuchi, Cedric Briones, Il-Hyung Lee","doi":"10.1021/acs.biochem.4c00483","DOIUrl":"https://doi.org/10.1021/acs.biochem.4c00483","url":null,"abstract":"Ubiquitylation is involved in various physiological processes, such as signaling and vesicle trafficking, whereas ubiquitin (UB) is considered an important clinical target. The polymeric addition of UB enables cargo molecules to be recognized specifically by multivalent binding interactions with UB-binding proteins, which results in various downstream processes. Recently, protein condensate formation by ubiquitylated proteins has been reported in many independent UB processes, suggesting its potential role in governing the spatial organization of ubiquitylated cargo proteins. We created modular polymeric UB binding motifs and polymeric UB cargos by synthetic bioconjugation and protein purification. Giant unilamellar vesicles with lipid raft composition were prepared to reconstitute the polymeric UB cargo organization on the membranes. Fluorescence imaging was used to observe the outcome. The polymeric UB cargos clustered on the membranes by forming a phase separation codomain during the interaction with the multivalent UB-binding conjugate. This phase separation was valence-dependent and strongly correlated with its potent ability to form protein condensate droplets in solution. Multivalent UB binding interactions exhibited a general trend toward the formation of phase-separated condensates and the resulting condensates were either in a liquid-like or solid-like state depending on the conditions and interactions. This suggests that the polymeric UB cargos on the plasma and endosomal membranes may use codomain phase separation to assist in the clustering of UB cargos on the membranes for cargo sorting. Our findings also indicate that such phase behavior model systems can be created by a modular synthetic approach that can potentially be used to further engineer biomimetic interactions in vitro.","PeriodicalId":28,"journal":{"name":"Biochemistry Biochemistry","volume":" ","pages":""},"PeriodicalIF":2.9,"publicationDate":"2025-02-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143497467","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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 Major Disease-Related Point Mutation in Spastin Dramatically Alters the Dynamics and Allostery of the Motor.

IF 2.9 3区生物学

Biochemistry Biochemistry Pub Date : 2025-02-26 DOI: 10.1021/acs.biochem.4c00693

Shehani Kahawatte, Amanda C Macke, Carter St Clair, Ruxandra I Dima

{"title":"A Major Disease-Related Point Mutation in Spastin Dramatically Alters the Dynamics and Allostery of the Motor.","authors":"Shehani Kahawatte, Amanda C Macke, Carter St Clair, Ruxandra I Dima","doi":"10.1021/acs.biochem.4c00693","DOIUrl":"https://doi.org/10.1021/acs.biochem.4c00693","url":null,"abstract":"Spastin is a microtubule-severing AAA+ ATPase that is highly expressed in neuronal cells and plays a crucial role in axonal growth, branching, and regeneration. This machine oligomerizes into hexamers in the presence of ATP and microtubule carboxy-terminal tails (CTTs). Conformational changes in spastin hexamers, powered by ATP hydrolysis, apply forces to the microtubule, ultimately leading to the severing of the filament. Mutations disrupt the normal function of spastin, impairing its ability to sever microtubules effectively and leading to abnormal microtubule dynamics in neurons characteristic of the set of neurodegenerative disorders called hereditary spastic paraplegias (HSP). Experimental studies have identified the HSP-related R591S (Drosophila melanogaster numbering) mutation as playing a crucial role in spastin. Given its significant role in HSP, we employed a combination of molecular dynamics simulations with machine learning and graph network-based approaches to identify and quantify the perturbations caused by the R591S HSP mutation on spastin's dynamics and allostery with functional implications. We found that the functional hexamer, upon HSP-related mutation, loses the ability to execute the primary motion associated with the severing action. The study of allosteric changes upon the mutation showed that the regions that are most perturbed are those involved in the formation of the interprotomer contacts. The mutation induces rigidity in the allosteric networks of the motor, making it more likely to experience loss of function as applied perturbations would not be easily dissipated by passing through a variety of alternative paths as in the wild-type (WT) spastin.","PeriodicalId":28,"journal":{"name":"Biochemistry Biochemistry","volume":" ","pages":""},"PeriodicalIF":2.9,"publicationDate":"2025-02-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143514001","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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

Disordered C-Terminus Plays a Critical Role in the Activity of the Small GTPase Ran.

IF 2.9 3区生物学

Biochemistry Biochemistry Pub Date : 2025-02-25 DOI: 10.1021/acs.biochem.4c00484

Wenyuan Wei, Melissa Valerio, Ning Ma, Hyunjun Kang, Le Xuan Truong Nguyen, Guido Marcucci, Nagarajan Vaidehi

{"title":"Disordered C-Terminus Plays a Critical Role in the Activity of the Small GTPase Ran.","authors":"Wenyuan Wei, Melissa Valerio, Ning Ma, Hyunjun Kang, Le Xuan Truong Nguyen, Guido Marcucci, Nagarajan Vaidehi","doi":"10.1021/acs.biochem.4c00484","DOIUrl":"https://doi.org/10.1021/acs.biochem.4c00484","url":null,"abstract":"Ran is a small GTPase of the Ras superfamily that governs nucleocytoplasmic transport, including that of miR-126, a microRNA that supports the homeostasis and expansion of leukemia stem cells (LSCs). Ran binds to Exportin 5 to facilitate the transport of precursor (pre)-miR-126 across the nuclear membrane for its maturation. Our goal is to inhibit Ran to prevent transport of pre-miR-126 to the cytoplasm. Like other Ras family proteins, targeting Ran with small molecules is challenging due to its relatively flat surface and lack of binding cavities. Ran's activity is regulated by a long and disordered C-terminus that provides opportunities for identifying cryptic binding pockets to target. We used a combination of molecular dynamics simulations and experiments and uncovered the critical role of the ensemble of the C-terminal conformations that enable the transition of Ran from the GTP-bound \"on state\" to its GDP-bound \"off-state\". We also showed that the Ran C-terminus allosterically modulates the conformations of residues in the nucleotide binding site and in the functionally relevant Switch 1 and 2 regions. Through computational deep mutational scans and experiments, we identified four residue hotspots L182, Y197, D200, and L201 at the core-C-terminus interface and four residue mutations V27A, E70D, N122A, and N122Y that mediate the allosteric communication between the core and switch regions. This information paves the way for our next step in the design of novel allosteric modulators for Ran.","PeriodicalId":28,"journal":{"name":"Biochemistry Biochemistry","volume":" ","pages":""},"PeriodicalIF":2.9,"publicationDate":"2025-02-25","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143497459","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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