Pub Date : 2025-12-04DOI: 10.1016/j.str.2025.11.008
Jakob R Riccabona,Johannes R Loeffler,Clara T Schoeder,Jens Meiler,Andrew B Ward,Monica Fernandez-Quintero
Deep-learning models have transformed structural biology by enabling reliable prediction of protein 3D structure models and providing confidence metrics such as predicted local distance difference test (pLDDT) to estimate local uncertainties. However, whether pLDDT reflects intrinsic protein flexibility remains unclear. Defining and quantifying flexibility and protein dynamics through experiments and computation is essential for advancing our ability to model and interpret conformational changes across different timescales.
{"title":"Rethinking what pLDDT really tells us about protein flexibility.","authors":"Jakob R Riccabona,Johannes R Loeffler,Clara T Schoeder,Jens Meiler,Andrew B Ward,Monica Fernandez-Quintero","doi":"10.1016/j.str.2025.11.008","DOIUrl":"https://doi.org/10.1016/j.str.2025.11.008","url":null,"abstract":"Deep-learning models have transformed structural biology by enabling reliable prediction of protein 3D structure models and providing confidence metrics such as predicted local distance difference test (pLDDT) to estimate local uncertainties. However, whether pLDDT reflects intrinsic protein flexibility remains unclear. Defining and quantifying flexibility and protein dynamics through experiments and computation is essential for advancing our ability to model and interpret conformational changes across different timescales.","PeriodicalId":22168,"journal":{"name":"Structure","volume":"1 1","pages":"2010-2012"},"PeriodicalIF":5.7,"publicationDate":"2025-12-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145680631","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}
Pub Date : 2025-12-04DOI: 10.1016/j.str.2025.11.001
Iktae Kim, Jeong-Yong Suh
In this issue of Structure, Henriques et al.1 present structural snapshots that capture distinct conformational states of the type I-F Cas1-Cas2/3 integrase complex, illustrating that foreign DNA binding triggers a large-scale domain rearrangement that enables prespacer delivery to the CRISPR array.
{"title":"Capture first, then deliver!","authors":"Iktae Kim, Jeong-Yong Suh","doi":"10.1016/j.str.2025.11.001","DOIUrl":"https://doi.org/10.1016/j.str.2025.11.001","url":null,"abstract":"<p><p>In this issue of Structure, Henriques et al.<sup>1</sup> present structural snapshots that capture distinct conformational states of the type I-F Cas1-Cas2/3 integrase complex, illustrating that foreign DNA binding triggers a large-scale domain rearrangement that enables prespacer delivery to the CRISPR array.</p>","PeriodicalId":22168,"journal":{"name":"Structure","volume":"33 12","pages":"2008-2009"},"PeriodicalIF":4.3,"publicationDate":"2025-12-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145688277","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}
Pub Date : 2025-12-04DOI: 10.1016/j.str.2025.11.007
Kate L White, Jin Young Kang, Renhong Yan, Katherine M Davis, Qianglin Fang, Lorena Saelices Gomez, Anthony W P Fitzpatrick, Mohammad T Mazhab-Jafari, Tatiana Galochkina
As 2025 comes to an end, we want to highlight some of the rising young faculty members who have published their exciting work from different areas of structural biology in Structure this year. We have asked them to tell us more about their interests, careers, and research programs.
{"title":"Our authors in 2025.","authors":"Kate L White, Jin Young Kang, Renhong Yan, Katherine M Davis, Qianglin Fang, Lorena Saelices Gomez, Anthony W P Fitzpatrick, Mohammad T Mazhab-Jafari, Tatiana Galochkina","doi":"10.1016/j.str.2025.11.007","DOIUrl":"https://doi.org/10.1016/j.str.2025.11.007","url":null,"abstract":"<p><p>As 2025 comes to an end, we want to highlight some of the rising young faculty members who have published their exciting work from different areas of structural biology in Structure this year. We have asked them to tell us more about their interests, careers, and research programs.</p>","PeriodicalId":22168,"journal":{"name":"Structure","volume":"33 12","pages":"1999-2007"},"PeriodicalIF":4.3,"publicationDate":"2025-12-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145688254","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}
Pub Date : 2025-11-28DOI: 10.1016/j.str.2025.11.006
Binning Sun, Jing Zheng, Yuan Fu, Fengyuan Tian, Hao Xiao, Su Li, Lingpeng Cheng, Ping Chen, Hongrong Liu
Podophage tails are too short to traverse the cell envelope and require internal core proteins to assemble into a transmembrane channel for genome delivery during infection. However, high-resolution structures of near-complete cores remain scarce. Here, we present the near-atomic-resolution cryo-electron microscopy (cryo-EM) structure of the drug-resistant E. coli phage E1004, which features a T7-like core-portal-tail structure with six P22-like tailspikes. We found that the cylindrical core comprises four proteins: gp17, gp27, gp28, and gp29. Gp29 forms a tetramer, while gp28 and gp27 assemble into octamers. Notably, there are sixteen copies of gp17 in two conformations, distinct from the small core protein gp6.7 in T7. The gp17-gp27 complex reveals the mechanism for mediating the symmetry adjustment at the core-portal interface. Moreover, comparative analysis with other podophage cores highlights diversity in core protein composition and organization, particularly among the small core proteins. We propose that these variations represent evolutionary adaptations to diverse host envelopes.
{"title":"Cryo-EM structure of drug-resistant Escherichia coli phage E1004 reveals a conserved cylindrical core among podophages","authors":"Binning Sun, Jing Zheng, Yuan Fu, Fengyuan Tian, Hao Xiao, Su Li, Lingpeng Cheng, Ping Chen, Hongrong Liu","doi":"10.1016/j.str.2025.11.006","DOIUrl":"https://doi.org/10.1016/j.str.2025.11.006","url":null,"abstract":"Podophage tails are too short to traverse the cell envelope and require internal core proteins to assemble into a transmembrane channel for genome delivery during infection. However, high-resolution structures of near-complete cores remain scarce. Here, we present the near-atomic-resolution cryo-electron microscopy (cryo-EM) structure of the drug-resistant <em>E. coli</em> phage E1004, which features a T7-like core-portal-tail structure with six P22-like tailspikes. We found that the cylindrical core comprises four proteins: gp17, gp27, gp28, and gp29. Gp29 forms a tetramer, while gp28 and gp27 assemble into octamers. Notably, there are sixteen copies of gp17 in two conformations, distinct from the small core protein gp6.7 in T7. The gp17-gp27 complex reveals the mechanism for mediating the symmetry adjustment at the core-portal interface. Moreover, comparative analysis with other podophage cores highlights diversity in core protein composition and organization, particularly among the small core proteins. We propose that these variations represent evolutionary adaptations to diverse host envelopes.","PeriodicalId":22168,"journal":{"name":"Structure","volume":"25 1","pages":""},"PeriodicalIF":5.7,"publicationDate":"2025-11-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145611929","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}
Pub Date : 2025-11-27DOI: 10.1016/j.str.2025.11.002
Ryan Ferrao, Jinjin Zhang, Chih-Chien Chou, Anthony Nieto, Derek Langeslay, Moon Chatterjee, Debi Jin, Magdeleine Hung, Ian Scott, Mark Nagel, Weimei Xing, Simon Letarte, Jenny Wang, Alexandre Ambrogelly, Eric B. Lansdon
Sacituzumab govitecan (SG) is a therapeutic antibody-drug conjugate globally approved for the treatment of breast cancer. SG targets the trophoblast cell-surface antigen-2 (Trop2) at the surface of cancer cells to deliver the cytotoxic topoisomerase I inhibitor SN-38 to the tumor microenvironment. SN-38 is covalently linked to the humanized monoclonal antibody (mAb) sacituzumab via a hydrolyzable linker. Here, we describe the 1.56-Å X-ray crystal structure and stoichiometry of the human Trop2 ectodomain in complex with a sacituzumab (hRS7) antigen-binding Fab fragment. The complex reveals a 2:2 stoichiometry where two sacituzumab Fabs bind across the two Trop2 dimer subunits, inducing a conformational change compared to the apo-structure. Cryo-electron microscopy (cryoEM) and size-exclusion chromatography in combination with multi-angle light scattering (SEC-MALS) analysis of the intact sacituzumab mAb bound to the Trop2 ECD reveals a complex whereby sacituzumab engages two Trop2 dimers in a 2:4 stoichiometry.
{"title":"The therapeutic antibody sacituzumab induces trophoblast cell-surface antigen-2 conformational rearrangement","authors":"Ryan Ferrao, Jinjin Zhang, Chih-Chien Chou, Anthony Nieto, Derek Langeslay, Moon Chatterjee, Debi Jin, Magdeleine Hung, Ian Scott, Mark Nagel, Weimei Xing, Simon Letarte, Jenny Wang, Alexandre Ambrogelly, Eric B. Lansdon","doi":"10.1016/j.str.2025.11.002","DOIUrl":"https://doi.org/10.1016/j.str.2025.11.002","url":null,"abstract":"Sacituzumab govitecan (SG) is a therapeutic antibody-drug conjugate globally approved for the treatment of breast cancer. SG targets the trophoblast cell-surface antigen-2 (Trop2) at the surface of cancer cells to deliver the cytotoxic topoisomerase I inhibitor SN-38 to the tumor microenvironment. SN-38 is covalently linked to the humanized monoclonal antibody (mAb) sacituzumab via a hydrolyzable linker. Here, we describe the 1.56-Å X-ray crystal structure and stoichiometry of the human Trop2 ectodomain in complex with a sacituzumab (hRS7) antigen-binding Fab fragment. The complex reveals a 2:2 stoichiometry where two sacituzumab Fabs bind across the two Trop2 dimer subunits, inducing a conformational change compared to the apo-structure. Cryo-electron microscopy (cryoEM) and size-exclusion chromatography in combination with multi-angle light scattering (SEC-MALS) analysis of the intact sacituzumab mAb bound to the Trop2 ECD reveals a complex whereby sacituzumab engages two Trop2 dimers in a 2:4 stoichiometry.","PeriodicalId":22168,"journal":{"name":"Structure","volume":"195 1","pages":""},"PeriodicalIF":5.7,"publicationDate":"2025-11-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145609740","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}
Pub Date : 2025-11-27DOI: 10.1016/j.str.2025.11.004
Ji Chen, Tao Li, Jiahua He, Sheng-You Huang
Accurate model building in intermediate-resolution cryo-EM maps normally requires flexible fitting of reliable initial structures. However, while deep learning-based methods such as AlphaFold2 can predict highly accurate structures, the predicted structures often differ from experimental EM maps on both global and local scales, which poses a great challenge to accurate model building in intermediate-resolution EM maps with such initial structures. Addressing the challenge, we propose CryoEvoBuild, an automated method for improved protein model building from intermediate-resolution EM maps through the effective integration of evolutionary and experimental information. CryoEvoBuild implements a novel domain-wise fitting, refinement, assembly, and rebuilding pipeline with a recycling framework guided by AlphaFold2. Extensive benchmarking on a diverse test set of 117 maps at 4.0–10.0 Å resolutions demonstrates that CryoEvoBuild significantly improves the accuracy of AF2-predicted structures and outperforms state-of-the-art approaches, including EMBuild and phenix.dock_and_rebuild.
{"title":"Protein model building for intermediate-resolution cryo-EM maps by integrating evolutionary and experimental information","authors":"Ji Chen, Tao Li, Jiahua He, Sheng-You Huang","doi":"10.1016/j.str.2025.11.004","DOIUrl":"https://doi.org/10.1016/j.str.2025.11.004","url":null,"abstract":"Accurate model building in intermediate-resolution cryo-EM maps normally requires flexible fitting of reliable initial structures. However, while deep learning-based methods such as AlphaFold2 can predict highly accurate structures, the predicted structures often differ from experimental EM maps on both global and local scales, which poses a great challenge to accurate model building in intermediate-resolution EM maps with such initial structures. Addressing the challenge, we propose CryoEvoBuild, an automated method for improved protein model building from intermediate-resolution EM maps through the effective integration of evolutionary and experimental information. CryoEvoBuild implements a novel domain-wise fitting, refinement, assembly, and rebuilding pipeline with a recycling framework guided by AlphaFold2. Extensive benchmarking on a diverse test set of 117 maps at 4.0–10.0 Å resolutions demonstrates that CryoEvoBuild significantly improves the accuracy of AF2-predicted structures and outperforms state-of-the-art approaches, including EMBuild and phenix.dock_and_rebuild.","PeriodicalId":22168,"journal":{"name":"Structure","volume":"9 1","pages":""},"PeriodicalIF":5.7,"publicationDate":"2025-11-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145609741","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}
Pub Date : 2025-11-27DOI: 10.1016/j.str.2025.11.003
Dieter Waschbüsch, Prosenjit Pal, Raja S. Nirujogi, Melanie Cavin, Jaijeet Singh, Dario R. Alessi, Amir R. Khan
Inherited mutations in VPS35 and LRRK2 kinase lead to hyperphosphorylation of Rab GTPases. RH2 domain-containing proteins from the RILP homology family, such as RILPL1, are Rab effectors that recognize the LRRK2-phosphorylated switch 2 threonine of phospho-Rab8A and phospho-Rab10. Phospho-Rabs are also seen on lysosomal membranes in complex with RILPL1 and TMEM55B, a 284-residue lysosomal membrane protein lacking homology to known proteins. Here, we report crystal structures of the cytosolic region 80–166 of TMEM55B alone and in complex with a C-terminal RILPL1 peptide, which we define as the TMEM55B-binding motif (TBM). The RILPL1 TBM sits in a shallow groove across two tandem RING-like domains of TMEM55B, each forming a Zn2+-stabilized 40-residue β-sandwich. Co-immunoprecipitation and mass spectrometry studies indicate that TMEM55B forms complexes independently of phospho-Rabs with conserved TBMs found in JIP3, JIP4, OCRL, WDR81, and TBC1D9B. These studies suggest that TMEM55B acts as a central hub for adaptor recruitment on lysosomes.
{"title":"Structural basis for binding of RILPL1 to TMEM55B reveals a lysosomal platform for adaptor assembly through a conserved peptide motif","authors":"Dieter Waschbüsch, Prosenjit Pal, Raja S. Nirujogi, Melanie Cavin, Jaijeet Singh, Dario R. Alessi, Amir R. Khan","doi":"10.1016/j.str.2025.11.003","DOIUrl":"https://doi.org/10.1016/j.str.2025.11.003","url":null,"abstract":"Inherited mutations in VPS35 and LRRK2 kinase lead to hyperphosphorylation of Rab GTPases. RH2 domain-containing proteins from the RILP homology family, such as RILPL1, are Rab effectors that recognize the LRRK2-phosphorylated switch 2 threonine of phospho-Rab8A and phospho-Rab10. Phospho-Rabs are also seen on lysosomal membranes in complex with RILPL1 and TMEM55B, a 284-residue lysosomal membrane protein lacking homology to known proteins. Here, we report crystal structures of the cytosolic region 80–166 of TMEM55B alone and in complex with a C-terminal RILPL1 peptide, which we define as the TMEM55B-binding motif (TBM). The RILPL1 TBM sits in a shallow groove across two tandem RING-like domains of TMEM55B, each forming a Zn<sup>2+</sup>-stabilized 40-residue β-sandwich. Co-immunoprecipitation and mass spectrometry studies indicate that TMEM55B forms complexes independently of phospho-Rabs with conserved TBMs found in JIP3, JIP4, OCRL, WDR81, and TBC1D9B. These studies suggest that TMEM55B acts as a central hub for adaptor recruitment on lysosomes.","PeriodicalId":22168,"journal":{"name":"Structure","volume":"21 1","pages":""},"PeriodicalIF":5.7,"publicationDate":"2025-11-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145609742","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}
Pub Date : 2025-11-27DOI: 10.1016/j.str.2025.11.005
Emily N. Bianchini, Carolina Pérez-Segura, Haitao Liu, Laura Luckenbaugh, John Flanagan, Yuanheng Cai, John Shanklin, Adam Zlotnick, Jodi A. Hadden-Perilla, Jianming Hu, Joseph C.-Y. Wang
HBV causes chronic infections that can lead to severe liver disease, yet current treatments rarely achieve a cure. The HBV capsid is a critical therapeutic target, but structural insights have largely relied on E. coli-derived particles lacking native modifications. Here, we present near-atomic resolution cryo-electron microscopy (EM) structures of HBV capsids purified from human embryonic kidney (HEK-293T) cells, capturing authentic architecture and post-translational modifications. A hydrophobic pocket at the intradimer interface harbors lipid-like densities corresponding to stearic and palmitic acids, confirmed by gas chromatography-mass spectrometry. Molecular dynamics simulations revealed that pocket accessibility is regulated by rotamer states of Lys96, Phe97, and Gln99, supporting an induced fit model of fatty acid binding. Reduced phosphorylation and increased RNA content further modulate capsid conformation and pocket openness. These findings highlight the dynamic regulation of HBV capsid structure and provide a framework for understanding how capsid conformational dynamics contribute to viral assembly and envelopment.
{"title":"Cryo-EM structures of HBV capsids from human cells at near-atomic resolution","authors":"Emily N. Bianchini, Carolina Pérez-Segura, Haitao Liu, Laura Luckenbaugh, John Flanagan, Yuanheng Cai, John Shanklin, Adam Zlotnick, Jodi A. Hadden-Perilla, Jianming Hu, Joseph C.-Y. Wang","doi":"10.1016/j.str.2025.11.005","DOIUrl":"https://doi.org/10.1016/j.str.2025.11.005","url":null,"abstract":"HBV causes chronic infections that can lead to severe liver disease, yet current treatments rarely achieve a cure. The HBV capsid is a critical therapeutic target, but structural insights have largely relied on <em>E. coli</em>-derived particles lacking native modifications. Here, we present near-atomic resolution cryo-electron microscopy (EM) structures of HBV capsids purified from human embryonic kidney (HEK-293T) cells, capturing authentic architecture and post-translational modifications. A hydrophobic pocket at the intradimer interface harbors lipid-like densities corresponding to stearic and palmitic acids, confirmed by gas chromatography-mass spectrometry. Molecular dynamics simulations revealed that pocket accessibility is regulated by rotamer states of Lys96, Phe97, and Gln99, supporting an induced fit model of fatty acid binding. Reduced phosphorylation and increased RNA content further modulate capsid conformation and pocket openness. These findings highlight the dynamic regulation of HBV capsid structure and provide a framework for understanding how capsid conformational dynamics contribute to viral assembly and envelopment.","PeriodicalId":22168,"journal":{"name":"Structure","volume":"71 1","pages":""},"PeriodicalIF":5.7,"publicationDate":"2025-11-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145609743","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}
Pub Date : 2025-11-24DOI: 10.1016/j.str.2025.10.001
Kazuki Obashi, Marie-Paule Strub, Justin W. Taraska
Ca2+-triggered exocytosis from neurons and endocrine cells is regulated by neuronal soluble N-ethylmaleimide-sensitive factor attachment receptor (SNARE) proteins. Conformational changes in syntaxin-1—the plasma membrane t-SNARE—are essential for vesicle docking and exocytosis. The nature of these conformational changes on the plasma membrane in living cells, however, remains largely unknown. Here, we develop a fluorescence system to map short-range conformational changes in syntaxin-1a in native plasma membranes of unroofed cells. We use a fluorescence resonance energy transfer (FRET) technique that employs site-specific protein labeling with unnatural fluorescent amino acids as donor fluorophores and colored transition metal ion acceptors bound to engineered di-histidine sites to map angstrom-scale distances. We find that phosphatidylinositol 4,5-bisphosphate (PIP2) regulates a conformational change in syntaxin-1a by modulating the structure of syntaxin-1a and its interaction with Munc18-1. Our results uncover new regulatory mechanisms of syntaxin-1a by PIP2 in the steps leading to Ca2+-triggered exocytosis.
{"title":"A PIP2-stabilized syntaxin-1a structure mapped with transition metal ion FRET and unnatural fluorescent amino acids at the plasma membrane","authors":"Kazuki Obashi, Marie-Paule Strub, Justin W. Taraska","doi":"10.1016/j.str.2025.10.001","DOIUrl":"https://doi.org/10.1016/j.str.2025.10.001","url":null,"abstract":"Ca<sup>2+</sup>-triggered exocytosis from neurons and endocrine cells is regulated by neuronal soluble N-ethylmaleimide-sensitive factor attachment receptor (SNARE) proteins. Conformational changes in syntaxin-1—the plasma membrane t-SNARE—are essential for vesicle docking and exocytosis. The nature of these conformational changes on the plasma membrane in living cells, however, remains largely unknown. Here, we develop a fluorescence system to map short-range conformational changes in syntaxin-1a in native plasma membranes of unroofed cells. We use a fluorescence resonance energy transfer (FRET) technique that employs site-specific protein labeling with unnatural fluorescent amino acids as donor fluorophores and colored transition metal ion acceptors bound to engineered di-histidine sites to map angstrom-scale distances. We find that phosphatidylinositol 4,5-bisphosphate (PIP2) regulates a conformational change in syntaxin-1a by modulating the structure of syntaxin-1a and its interaction with Munc18-1. Our results uncover new regulatory mechanisms of syntaxin-1a by PIP2 in the steps leading to Ca<sup>2+</sup>-triggered exocytosis.","PeriodicalId":22168,"journal":{"name":"Structure","volume":"167 1","pages":""},"PeriodicalIF":5.7,"publicationDate":"2025-11-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145583887","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}
Muscle-specific receptor tyrosine kinase (MuSK) is a single-pass transmembrane protein expressed on skeletal muscle. MuSK is activated by binding of nerve-derived agrin with the help of muscle coreceptor LRP4, leading to the clustering of acetylcholine receptors (AChR), which is required for the formation and maintenance of functional neuromuscular junctions. The structural mechanism of MuSK activation by physiological and artificial agonistic agents has remained elusive. In this study, we isolated a 27-residue linear peptide (L1) that binds human MuSK with high affinity. Genetic fusion of L1 to either the N or C termini of the human IgG Fc resulted in two different versions of MuSK dimerizers, denoted as L1-Fc and Fc-L1. Only Fc-L1 activated MuSK on myotubes and induced AChR clustering. Crystallographic analysis of MuSK-L1 interactions revealed that MuSK activation requires a particular dimeric conformation, pointing toward the importance of the lateral size of the receptor complex at the muscle cell surface.
{"title":"Muscle-specific tyrosine kinase activation by a peptide-based dimerizer is orientation dependent","authors":"Fumiya Mizutani, Kyoko Matoba, Hayden Peacock, Mitsuhiro Yamada, Emiko Mihara, Osamu Higuchi, Hiroaki Suga, Takao Arimori, Junichi Takagi","doi":"10.1016/j.str.2025.10.018","DOIUrl":"https://doi.org/10.1016/j.str.2025.10.018","url":null,"abstract":"Muscle-specific receptor tyrosine kinase (MuSK) is a single-pass transmembrane protein expressed on skeletal muscle. MuSK is activated by binding of nerve-derived agrin with the help of muscle coreceptor LRP4, leading to the clustering of acetylcholine receptors (AChR), which is required for the formation and maintenance of functional neuromuscular junctions. The structural mechanism of MuSK activation by physiological and artificial agonistic agents has remained elusive. In this study, we isolated a 27-residue linear peptide (L1) that binds human MuSK with high affinity. Genetic fusion of L1 to either the N or C termini of the human IgG Fc resulted in two different versions of MuSK dimerizers, denoted as L1-Fc and Fc-L1. Only Fc-L1 activated MuSK on myotubes and induced AChR clustering. Crystallographic analysis of MuSK-L1 interactions revealed that MuSK activation requires a particular dimeric conformation, pointing toward the importance of the lateral size of the receptor complex at the muscle cell surface.","PeriodicalId":22168,"journal":{"name":"Structure","volume":"223 1","pages":""},"PeriodicalIF":5.7,"publicationDate":"2025-11-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"145583815","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}
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