Oakley Streeter, Ke Shi, Joseph Vavra, Hideki Aihara, James M Ervasti, Robert Evans, Joseph M Muretta
{"title":"Human dystrophin tandem calponin homology actin-binding domain crystallized in a closed-state conformation.","authors":"Oakley Streeter, Ke Shi, Joseph Vavra, Hideki Aihara, James M Ervasti, Robert Evans, Joseph M Muretta","doi":"10.1107/S2059798325001457","DOIUrl":"10.1107/S2059798325001457","url":null,"abstract":"<p><p>The structure of the N-terminal actin-binding domain of human dystrophin was determined at 1.94 Å resolution. Each chain in the asymmetric unit exists in a `closed' conformation, with the first and second calponin homology (CH) domains directly interacting via a 2500.6 Å<sup>2</sup> interface. The positioning of the individual CH domains is comparable to the domain-swapped dimer seen in previous human dystrophin and utrophin actin-binding domain 1 structures. The CH1 domain is highly similar to the actin-bound utrophin structure and structural homology suggests that the `closed' single-chain conformation opens during actin binding to mitigate steric clashes between CH2 and actin.</p>","PeriodicalId":7116,"journal":{"name":"Acta Crystallographica. Section D, Structural Biology","volume":" ","pages":"122-129"},"PeriodicalIF":2.6,"publicationDate":"2025-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11883666/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143497639","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Alexandra Males, Olga V Moroz, Elena Blagova, Astrid Munch, Gustav H Hansen, Annette H Johansen, Lars H Østergaard, Dorotea R Segura, Alexander Eddenden, Anne V Due, Martin Gudmand, Jesper Salomon, Sebastian R Sørensen, João Paulo L Franco Cairo, Mark Nitz, Roland A Pache, Rebecca M Vejborg, Sandeep Bhosale, David J Vocadlo, Gideon J Davies, Keith S Wilson
{"title":"Expansion of the diversity of dispersin scaffolds.","authors":"Alexandra Males, Olga V Moroz, Elena Blagova, Astrid Munch, Gustav H Hansen, Annette H Johansen, Lars H Østergaard, Dorotea R Segura, Alexander Eddenden, Anne V Due, Martin Gudmand, Jesper Salomon, Sebastian R Sørensen, João Paulo L Franco Cairo, Mark Nitz, Roland A Pache, Rebecca M Vejborg, Sandeep Bhosale, David J Vocadlo, Gideon J Davies, Keith S Wilson","doi":"10.1107/S205979832500110X","DOIUrl":"10.1107/S205979832500110X","url":null,"abstract":"<p><p>Microorganisms are known to secrete copious amounts of extracellular polymeric substances (EPS) that form complex matrices around the cells to shield them against external stresses, to maintain structural integrity and to influence their environment. Many microorganisms also secrete enzymes that are capable of remodelling or degrading EPS in response to various environmental cues. One key enzyme class is the poly-β-1,6-linked N-acetyl-D-glucosamine (PNAG)-degrading glycoside hydrolases, of which the canonical member is dispersin B (DspB) from CAZy family GH20. We sought to test the hypothesis that PNAG-degrading enzymes would be present across family GH20, resulting in expansion of the sequence and structural space and thus the availability of PNAGases. Phylogenetic analysis revealed that several microorganisms contain potential DspB-like enzymes. Six of these were expressed and characterized, and four crystal structures were determined (two of which were in complex with the established GH20 inhibitor 6-acetamido-6-deoxy-castanospermine and one with a bespoke disaccharide β-1,6-linked thiazoline inhibitor). One enzyme expressed rather poorly, which restricted crystal screening and did not allow activity measurements. Using synthetic PNAG oligomers and MALDI-TOF analysis, two of the five enzymes tested showed preferential endo hydrolytic activity. Their sequences, having only 26% identity to the pioneer enzyme DspB, highlight the considerable array of previously unconsidered dispersins in nature, greatly expanding the range of potential dispersin backbones available for societal application and engineering.</p>","PeriodicalId":7116,"journal":{"name":"Acta Crystallographica. Section D, Structural Biology","volume":" ","pages":"130-146"},"PeriodicalIF":2.6,"publicationDate":"2025-03-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11883664/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143522423","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Has AlphaFold3 achieved success for RNA?","authors":"Clément Bernard, Guillaume Postic, Sahar Ghannay, Fariza Tahi","doi":"10.1107/S2059798325000592","DOIUrl":"10.1107/S2059798325000592","url":null,"abstract":"<p><p>Predicting the 3D structure of RNA is a significant challenge despite ongoing advancements in the field. Although AlphaFold has successfully addressed this problem for proteins, RNA structure prediction raises difficulties due to the fundamental differences between proteins and RNA, which hinder its direct adaptation. The latest release of AlphaFold, AlphaFold3, has broadened its scope to include multiple different molecules such as DNA, ligands and RNA. While the AlphaFold3 article discussed the results for the last CASP-RNA data set, the scope of its performance and the limitations for RNA are unclear. In this article, we provide a comprehensive analysis of the performance of AlphaFold3 in the prediction of 3D structures of RNA. Through an extensive benchmark over five different test sets, we discuss the performance and limitations of AlphaFold3. We also compare its performance with ten existing state-of-the-art ab initio, template-based and deep-learning approaches. Our results are freely available on the EvryRNA platform at https://evryrna.ibisc.univ-evry.fr/evryrna/alphafold3/.</p>","PeriodicalId":7116,"journal":{"name":"Acta Crystallographica. Section D, Structural Biology","volume":" ","pages":"49-62"},"PeriodicalIF":2.6,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11804252/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143045244","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Charles S Bond, Paul M G Curmi, John R Helliwell, Alan Riboldi-Tunnicliffe, Rachel M Williamson
{"title":"Stephen Harrop (1966-2024).","authors":"Charles S Bond, Paul M G Curmi, John R Helliwell, Alan Riboldi-Tunnicliffe, Rachel M Williamson","doi":"10.1107/S2059798325000890","DOIUrl":"10.1107/S2059798325000890","url":null,"abstract":"<p><p>Stephen Harrop is remembered.</p>","PeriodicalId":7116,"journal":{"name":"Acta Crystallographica. Section D, Structural Biology","volume":" ","pages":"85-88"},"PeriodicalIF":2.6,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143078132","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Reconsideration of the P-clusters in VFe proteins using the bond-valence method: towards their electron transfer and protonation.","authors":"Zhen Lang Xie, Wan Ting Jin, Zhao Hui Zhou","doi":"10.1107/S2059798325000415","DOIUrl":"10.1107/S2059798325000415","url":null,"abstract":"<p><p>P-clusters have been statistically analysed using the bond-valence sum (BVS) method together with weighting schemes. The crystallographic data come from the VFe proteins deposited in the Protein Data Bank (PDB) with high resolutions of better than 1.35 Å. Calculations show that the formal oxidation state of a P<sup>1+</sup> cluster can be assigned as 2Fe<sup>3+</sup>6Fe<sup>2+</sup> with high electron delocalization, giving the same oxidation state as that of P<sup>N</sup> clusters in VFe proteins. Further comprehensive comparisons of the bond distances suggest that the hydroxyl groups of the β-153 serine residues in P<sup>1+</sup> and P<sup>N</sup> clusters are in the protonated state, where the Fe6 atoms have the same oxidation state as Fe<sup>2+</sup>. During the transition from P<sup>N</sup> to P<sup>1+</sup>, cleavage of the Fe6-S1 bond is accompanied by the formation of a weak coordination between the Fe6 atom and the hydroxyl group of the β-153 serine residue in the P<sup>1+</sup> cluster of the VFe protein. Similarly, oxidation of P<sup>N</sup> to P<sup>1+</sup>/P<sup>2+</sup> clusters corresponds to the coordination of Fe6(II) by the hydroxyl group of the β-188 serine residue and of Fe5(II) by the peptide amine group of the α-88 cysteine residue in the MoFe protein of Azotobacter vinelandiis without electron and proton transfers.</p>","PeriodicalId":7116,"journal":{"name":"Acta Crystallographica. Section D, Structural Biology","volume":" ","pages":"77-84"},"PeriodicalIF":2.6,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142998340","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Pranav N M Shah, Ruben Sanchez-Garcia, David I Stuart
{"title":"TomoCPT: a generalizable model for 3D particle detection and localization in cryo-electron tomograms.","authors":"Pranav N M Shah, Ruben Sanchez-Garcia, David I Stuart","doi":"10.1107/S2059798325000865","DOIUrl":"10.1107/S2059798325000865","url":null,"abstract":"<p><p>Cryo-electron tomography is a rapidly developing field for studying macromolecular complexes in their native environments and has the potential to revolutionize our understanding of protein function. However, fast and accurate identification of particles in cryo-tomograms is challenging and represents a significant bottleneck in downstream processes such as subtomogram averaging. Here, we present tomoCPT (Tomogram Centroid Prediction Tool), a transformer-based solution that reformulates particle detection as a centroid-prediction task using Gaussian labels. Our approach, which is built upon the SwinUNETR architecture, demonstrates superior performance compared with both conventional binary labelling strategies and template matching. We show that tomoCPT effectively generalizes to novel particle types through zero-shot inference and can be significantly enhanced through fine-tuning with limited data. The efficacy of tomoCPT is validated using three case studies: apoferritin, achieving a resolution of 3.0 Å compared with 3.3 Å using template matching, SARS-CoV-2 spike proteins on cell surfaces, yielding an 18.3 Å resolution map where template matching proved unsuccessful, and rubisco molecules within carboxysomes, reaching 8.0 Å resolution. These results demonstrate the ability of tomoCPT to handle varied scenarios, including densely packed environments and membrane-bound proteins. The implementation of the tool as a command-line program, coupled with its minimal data requirements for fine-tuning, makes it a practical solution for high-throughput cryo-ET data-processing workflows.</p>","PeriodicalId":7116,"journal":{"name":"Acta Crystallographica. Section D, Structural Biology","volume":" ","pages":"63-76"},"PeriodicalIF":2.6,"publicationDate":"2025-02-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11804251/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143187910","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"AlphaFold-guided molecular replacement for solving challenging crystal structures.","authors":"Wei Wang, Zhen Gong, Wayne A Hendrickson","doi":"10.1107/S2059798324011999","DOIUrl":"10.1107/S2059798324011999","url":null,"abstract":"<p><p>Molecular replacement (MR) is highly effective for biomolecular crystal structure determination, increasingly so as the database of known structures has increased. For candidates without recognizable similarity to known structures, however, crystal structure analyses have nearly always required experiments for de novo phase evaluation. Now, with the unprecedented accuracy of AlphaFold predictions of protein structures from amino-acid sequences, an appreciable expansion of the reach of MR for proteins is realized. Here, we sought to automate an AlphaFold-guided MR procedure that tailors predictions to the MR problem at hand. We first optimized the reliability cutoff parameters for residue inclusion as tested in application to a previously MR-intractable problem. We then examined cases where AlphaFold by default predicts a conformation alternative to that of the candidate structure, devising tests for MR solution either from domain-specific predictions or from predictions based on diverse sequence subclusters. We tested subclustering procedures on an enzyme system that entails multiple MR-challenging conformations. The overall process as implemented in Phenix automatically surveys a succession of trials of increasing computational complexity until an MR solution is found or the options are exhausted. Validated MR solutions were found for 92% of one set of 158 challenging problems from the PDB and 93% of those from a second set of 215 challenges. Thus, many crystal structure analyses that previously required experimental phase evaluation can now be solved by AlphaFold-guided MR. In effect, this and related MR approaches are de novo phasing methods.</p>","PeriodicalId":7116,"journal":{"name":"Acta Crystallographica. Section D, Structural Biology","volume":" ","pages":"4-21"},"PeriodicalIF":2.6,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11740581/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142875864","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
Christoph Mueller-Dieckmann, Anna J Warren, David G Waterman
{"title":"Making the most of an abundance of data.","authors":"Christoph Mueller-Dieckmann, Anna J Warren, David G Waterman","doi":"10.1107/S205979832401204X","DOIUrl":"10.1107/S205979832401204X","url":null,"abstract":"<p><p>The Guest Editors introduce the special issue based on talks at the CCP4 Study Weekend 2023. The virtual issue is available at https://journals.iucr.org/special_issues/2024/CCP42023/.</p>","PeriodicalId":7116,"journal":{"name":"Acta Crystallographica. Section D, Structural Biology","volume":" ","pages":"1-3"},"PeriodicalIF":2.6,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11740582/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142863008","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"Useful experimental aspects of small-wedge synchrotron crystallography for accurate structure analysis of protein molecules.","authors":"Kunio Hirata","doi":"10.1107/S2059798324011987","DOIUrl":"10.1107/S2059798324011987","url":null,"abstract":"<p><p>Recent advances in low-emittance synchrotron X-ray technology and highly sensitive photon-counting detectors have revolutionized protein micro-crystallography in structural biology. These developments and improvements to sample-exchange robots and beamline control have paved the way for automated and efficient unattended data collection. This study analyzed protein crystal structures such as type 2 angiotensin II receptor, CNNM/CorC membrane proteins and polyhedral protein crystals using small-wedge synchrotron crystallography (SWSX), which dramatically improves measurement efficiency through automated measurement. We evaluated the data quality using SWSX, focusing on `massive data collection'. In this context, `massive' refers to data sets with a multiplicity exceeding 100. The findings could potentially lead to the development of more efficient experimental conditions, such as obtaining high-resolution data using a smaller number of crystals. We have demonstrated that the application of machine learning, a modern key component of data science, to classify data groups is an integral part of the analysis process and may play a crucial role in improving data quality. These results indicate that SWSX is one of the essential candidates for crystal structure analysis methods for difficult-to-analyze samples: it can enable diverse and complex protein functional analysis.</p>","PeriodicalId":7116,"journal":{"name":"Acta Crystallographica. Section D, Structural Biology","volume":" ","pages":"22-37"},"PeriodicalIF":2.6,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11740584/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142881067","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
{"title":"The 1.3 Å resolution structure of the truncated group Ia type IV pilin from Pseudomonas aeruginosa strain P1.","authors":"Nicholas Bragagnolo, Gerald F Audette","doi":"10.1107/S205979832401132X","DOIUrl":"10.1107/S205979832401132X","url":null,"abstract":"<p><p>The type IV pilus is a diverse molecular machine capable of conferring a variety of functions and is produced by a wide range of bacterial species. The ability of the pilus to perform host-cell adherence makes it a viable target for the development of vaccines against infection by human pathogens such as Pseudomonas aeruginosa. Here, the 1.3 Å resolution crystal structure of the N-terminally truncated type IV pilin from P. aeruginosa strain P1 (ΔP1) is reported, the first structure of its phylogenetically linked group (group I) to be discussed in the literature. The structure was solved from X-ray diffraction data that were collected 20 years ago with a molecular-replacement search model generated using AlphaFold; the effectiveness of other search models was analyzed. Examination of the high-resolution ΔP1 structure revealed a solvent network that aids in maintaining the fold of the protein. On comparing the sequence and structure of P1 with a variety of type IV pilins, it was observed that there are cases of higher structural similarities between the phylogenetic groups of P. aeruginosa than there are between the same phylogenetic group, indicating that a structural grouping of pilins may be necessary in developing antivirulence drugs and vaccines. These analyses also identified the α-β loop as the most structurally diverse domain of the pilins, which could allow it to serve a role in pilus recognition. Studies of ΔP1 in vitro polymerization demonstrate that the optimal hydrophobic catalyst for the oligomerization of the pilus from strain K122 is not conducive for pilus formation of ΔP1; a model of a three-start helical assembly using the ΔP1 structure indicates that the α-β loop and the D-loop prevent in vitro polymerization.</p>","PeriodicalId":7116,"journal":{"name":"Acta Crystallographica. Section D, Structural Biology","volume":" ","pages":"834-849"},"PeriodicalIF":2.6,"publicationDate":"2024-12-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC11626772/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142749698","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}