Journal of Molecular Biology最新文献

{"title":"The Evolution of Sequence Specificity in a DNA Binding Protein Family","authors":"Meghna Nandy ,&nbsp;Madhumitha Krishnaswamy ,&nbsp;Mohak Sharda ,&nbsp;Aswin Sai Narain Seshasayee","doi":"10.1016/j.jmb.2025.169177","DOIUrl":"10.1016/j.jmb.2025.169177","url":null,"abstract":"<div><div>Transcriptional regulation enables bacteria to adjust to its environment. This is driven by transcription factors (TFs), which display DNA site recognition specificity with some flexibility built in. TFs, however, are not considered essential to a minimal cellular life. How did they evolve? It has been hypothesized that TFs evolve by gaining specificity (and other functions) on a background of non-specific chromosome structuring proteins. We used the IHF/HU family of DNA binding proteins, in which IHF binds DNA in a sequence-specific manner, whereas HU binds more indiscriminately, to test this hypothesis. We show that HUβ has been present from the bacterial root, while both IHF subunits emerged much later and diversified in <em>Proteobacteria</em>, with HUα having possibly arisen from transfer events in <em>Gammaproteobacteria</em>. By reconstructing ancestral sequences <em>in-silico</em> on a rooted phylogeny of IHF/HU we show that the common ancestor of this family was probably HU-like and therefore non-specific in binding DNA. IHF evolved from a branch of HU after HU had substantially diverged. Various residues characteristic of IHFα and shown to be involved in specific sequence recognition (at least in <em>E. coli</em>) have likely been co-opted from preexisting residues in HU, while those residues of IHFβ have likely evolved independently, suggesting that each of the IHF subunits has undergone different trajectories to acquire their DNA binding properties.</div></div>","PeriodicalId":369,"journal":{"name":"Journal of Molecular Biology","volume":"437 14","pages":"Article 169177"},"PeriodicalIF":4.7,"publicationDate":"2025-04-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143924349","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}

{"title":"Are Allosteric Mechanisms Conserved Among Homologues?","authors":"Aron W Fenton, Zoe A Hoffpauir, Tyler A Martin, Robert A Harris, Audrey L Lamb","doi":"10.1016/j.jmb.2025.169176","DOIUrl":"https://doi.org/10.1016/j.jmb.2025.169176","url":null,"abstract":"<p><p>Conservation of allosteric mechanisms among homologues is often assumed but seldom tested. This assumption underpins key concepts like coevolution of residues involved in allosteric mechanisms and the comparison of structures of two different homologues to gain insights into allosteric mechanisms. As an initial assessment of whether allosteric mechanisms are conserved among homologues, this work reviews what is known about the allosteric mechanisms of liver pyruvate kinase (LPYK) vs. skeletal muscle pyruvate kinase (M₁PYK), framed within a two-ligand allosteric energy cycle description of allosteric regulation. Selective observations from other PYK homologues are included when relevant. The primary focus of this review is on functional data, while expressing caution regarding the interpretation of allosteric mechanisms based solely on available X-ray crystallographic structures. Additionally, this review considers types of data that are currently lacking for these two PYK homologues, highlighting potential techniques that could be valuable for evaluating the conservation of allosteric mechanisms among homologues. In particular, a hybrid tetramer technique that has been used to study bacterial phosphofructokinases 1 is summarized. Interestingly, despite a high degree of similarity (66.5% sequence identity) between the LPYK and rM₁PYK proteins, the available functional comparisons do not provide strong evidence for conserved allosteric mechanisms. Lastly, we consider whether insights into native allosteric mechanisms are relevant to allosteric mechanisms associated with allosteric drug designs.</p>","PeriodicalId":369,"journal":{"name":"Journal of Molecular Biology","volume":" ","pages":"169176"},"PeriodicalIF":4.7,"publicationDate":"2025-04-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143951325","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}

{"title":"Dynamic Allostery: Evolution's Double-Edged Sword in Protein Function and Disease.","authors":"Paul Campitelli, I Can Kazan, Sean Hamilton, S Banu Ozkan","doi":"10.1016/j.jmb.2025.169175","DOIUrl":"https://doi.org/10.1016/j.jmb.2025.169175","url":null,"abstract":"<p><p>Allostery is a core mechanism in biology that allows proteins to communicate and regulate activity over long structural distances. While classical models of allostery focus on conformational changes triggered by ligand binding, dynamic allostery-where protein function is modulated through alterations in thermal fluctuations without major conformational shifts-has emerged as a critical evolutionary mechanism. This review explores how evolution leverages dynamic allostery to fine-tune protein function through subtle mutations at distal sites, preserving core structural architecture while dramatically altering functional properties. Using a combination of computational approaches including Dynamic Flexibility Index (DFI), Dynamic Coupling Index (DCI), and vibrational density of states (VDOS) analysis, we demonstrate that functional adaptations in proteins often involve \"hinge-shift\" mechanisms, where redistribution of rigid and flexible regions modulates collective motions without changing the overall fold. This evolutionary principle is a double-edged sword: the same mechanisms that enable functional innovation also create vulnerabilities that can be exploited in disease states. Disease-associated variants frequently occur at positions highly coupled to functional sites despite being physically distant, forming Dynamic Allosteric Residue Couples (DARC sites). We demonstrate applications of these principles in understanding viral evolution, drug resistance, and capsid assembly dynamics. Understanding dynamic allostery provides critical insights into protein evolution and offers new avenues for therapeutic interventions targeting allosteric regulation.</p>","PeriodicalId":369,"journal":{"name":"Journal of Molecular Biology","volume":" ","pages":"169175"},"PeriodicalIF":4.7,"publicationDate":"2025-04-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143951799","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}

{"title":"Science in the U.S. Is Under Attack – What Can We Do?","authors":"","doi":"10.1016/j.jmb.2025.169165","DOIUrl":"10.1016/j.jmb.2025.169165","url":null,"abstract":"","PeriodicalId":369,"journal":{"name":"Journal of Molecular Biology","volume":"437 14","pages":"Article 169165"},"PeriodicalIF":4.7,"publicationDate":"2025-04-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143886276","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}

{"title":"GPCRchimeraDB: A Database of Chimeric G Protein-Coupled Receptors (GPCRs) to Assist Their Design","authors":"Charlotte Crauwels ,&nbsp;Adrián Díaz ,&nbsp;Wim Vranken","doi":"10.1016/j.jmb.2025.169164","DOIUrl":"10.1016/j.jmb.2025.169164","url":null,"abstract":"<div><div>G protein-coupled receptors (GPCRs) are membrane proteins crucial to numerous diseases, yet many remain poorly characterized and untargeted by drugs. Chimeric GPCRs have emerged as valuable tools for elucidating GPCR function by facilitating the identification of signaling pathways, resolving structures, and discovering novel ligands of poorly understood GPCRs. Such chimeric GPCRs are obtained by merging a well- and less-well-characterized GPCR at the intracellular limits of their transmembrane regions or intracellular loops, leveraging knowledge transfer from the well-characterized GPCR. However, despite the engineering of over 200 chimeric GPCRs to date, the design process remains largely trial-and-error and lacks a standardized approach. To address this gap, we introduce GPCRchimeraDB (<span><span>https://www.bio2byte.be/gpcrchimeradb/</span><svg><path></path></svg></span>), the first comprehensive database dedicated to chimeric GPCRs. It catalogs 212 chimeric receptors, identified through literature review, and includes 1,755 class A natural GPCRs, enabling connections between chimeras and their parent receptors while facilitating the exploration of novel parent combinations. Both chimeric and natural GPCR entries are extensively described at the sequence, structural, and biophysical level through a range of visualization tools, with annotations from resources like UniProt and GPCRdb and predictions from AlphaFold2 and b2btools. Additionally, GPCRchimeraDB offers a GPCR sequence aligner and a feature comparator to investigate differences between natural and chimeric receptors. It also provides design guidelines to support rational chimera engineering. GPCRchimeraDB is therefore a resource to facilitate and optimize the design of new chimeras, so helping to gain insights into poorly characterized receptors and contributing to advances in GPCR therapeutic development.</div></div>","PeriodicalId":369,"journal":{"name":"Journal of Molecular Biology","volume":"437 14","pages":"Article 169164"},"PeriodicalIF":4.7,"publicationDate":"2025-04-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143908290","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}

{"title":"CIWARS: A Web Server for Antibiotic Resistance Surveillance Using Longitudinal Metagenomic Data","authors":"Muhit Islam Emon ,&nbsp;Yat Fei Cheung ,&nbsp;James Stoll ,&nbsp;Monjura Afrin Rumi ,&nbsp;Connor Brown ,&nbsp;Joung Min Choi ,&nbsp;Nazifa Ahmed Moumi ,&nbsp;Shafayat Ahmed ,&nbsp;Haoqiu Song ,&nbsp;Justin Sein ,&nbsp;Shunyu Yao ,&nbsp;Ahmad Khan ,&nbsp;Suraj Gupta ,&nbsp;Rutwik Kulkarni ,&nbsp;Ali Butt ,&nbsp;Peter Vikesland ,&nbsp;Amy Pruden ,&nbsp;Liqing Zhang","doi":"10.1016/j.jmb.2025.169159","DOIUrl":"10.1016/j.jmb.2025.169159","url":null,"abstract":"<div><div>The rise of antibiotic resistance (AR) poses a substantial threat to human and animal health, food security, and economic stability. Wastewater-based surveillance (WBS) has emerged as a powerful strategy for population-level AR monitoring, providing valuable data to guide public health and policy decisions. Metagenomic sequencing is especially promising, as it can yield comprehensive profiles of antibiotic resistance genes (ARGs) and other genes relevant to AR in a single run. However, online analytical platforms to facilitate analysis of longitudinal metagenomic data are lacking. To address this, we introduce <strong>C</strong>yber<strong>I</strong>nfrastructure for <strong>W</strong>aterborne <strong>A</strong>ntibiotic <strong>R</strong>esistance <strong>S</strong>urveillance (CIWARS), a web server configured for characterizing key AR trends from longitudinal metagenomic WBS data. CIWARS offers comprehensive profiling of ARGs and taxonomic profiling of pathogen-associated bacterial taxonomic groups, identifies potential associations of ARGs with mobile genetic elements (MGEs) and pathogen-containing taxa, and assesses resistome risk based on the co-occurrence of ARGs, MGEs, and pathogen-like sequences. Additionally, it detects anomalous AR indicators over time, aiding in identifying potential events of concern, such as the emergence of resistant strains or outbreaks. Through interactive temporal data visualization, CIWARS enables AR monitoring and can serve as a tool to inform effective and timely interventions to mitigate the spread and transmission of AR. Here, CIWARS is demonstrated using longitudinal metagenomic data from a wastewater treatment plant (WWTP) influent and effluent, but it can be extended to any environment. CIWARS provides a valuable tool to support global efforts to combat the evolution and spread of AR, while also guiding agricultural and public health efforts aimed at optimizing antibiotic use. The web server is freely available at <span><span>https://ciwars.cs.vt.edu/</span><svg><path></path></svg></span>.</div></div>","PeriodicalId":369,"journal":{"name":"Journal of Molecular Biology","volume":"437 15","pages":"Article 169159"},"PeriodicalIF":4.7,"publicationDate":"2025-04-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144053465","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}

{"title":"Mitophagy in Neurons: Mechanisms Regulating Mitochondrial Turnover and Neuronal Homeostasis.","authors":"Bishal Basak, Erika L F Holzbaur","doi":"10.1016/j.jmb.2025.169161","DOIUrl":"https://doi.org/10.1016/j.jmb.2025.169161","url":null,"abstract":"<p><p>Mitochondrial quality control is instrumental in regulating neuronal health and survival. The receptor-mediated clearance of damaged mitochondria by autophagy, known as mitophagy, plays a key role in controlling mitochondrial homeostasis. Mutations in genes that regulate mitophagy are causative for familial forms of neurological disorders including Parkinson's disease (PD) and Amyotrophic lateral sclerosis (ALS). PINK1/Parkin-dependent mitophagy is the best studied mitophagy pathway, while more recent work has brought to light additional mitochondrial quality control mechanisms that operate either in parallel to or independent of PINK1/Parkin mitophagy. Here, we discuss our current understanding of mitophagy mechanisms operating in neurons to govern mitochondrial homeostasis. We also summarize progress in our understanding of the links between mitophagic dysfunction and neurodegeneration, and highlight the potential for therapeutic interventions to maintain mitochondrial health and neuronal function.</p>","PeriodicalId":369,"journal":{"name":"Journal of Molecular Biology","volume":" ","pages":"169161"},"PeriodicalIF":4.7,"publicationDate":"2025-04-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143958456","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}

{"title":"Switch II Pocket Inhibitor Allosterically Freezes KRASG12D Nucleotide-binding Site and Arrests the GTPase Cycle","authors":"Ha-Neul Kim ,&nbsp;Geneviève M.C. Gasmi-Seabrook ,&nbsp;Arisa Uchida ,&nbsp;Teklab Gebregiworgis ,&nbsp;Christopher B. Marshall ,&nbsp;Mitsuhiko Ikura","doi":"10.1016/j.jmb.2025.169162","DOIUrl":"10.1016/j.jmb.2025.169162","url":null,"abstract":"<div><div><em>KRAS</em> is frequently mutated in multiple cancers, with the most common mutation being G12D. The recently developed KRAS^G12D inhibitor MRTX1133 binds a cryptic allosteric pocket near switch II (SII-P), similar to covalent G12C inhibitors, with remarkable picoM non-covalent affinity. Despite its advancement to clinical trials, some aspects of the molecular mechanisms-of-action remain unclear, indicating a need to uncover the mechanisms underlying MRTX1133 efficacy and potential acquired resistance, thus we characterized the biochemical and biophysical outcomes of MRTX1133 binding KRAS. Hydrogen/deuterium exchange experiments showed that MRTX1133 binding to the induced SII-P reduces the overall conformational plasticity of KRAS^G12D. This extends well beyond SII-P, with the nucleotide-binding regions (P-loop and G-3/4/5-box motifs) particularly exhibiting stabilization. This conformational rigidification by MRTX1133 is coupled with complete arrest of the GTPase cycle: When the compound engages KRAS^G12D-GDP, both intrinsic and GEF-mediated nucleotide exchange are blocked while engagement of KRAS^G12D-GTP blocks both intrinsic and GAP-mediated hydrolysis. MRTX1133 attenuates the interaction between activated KRAS^G12D and the RAS-binding domain of the effector BRAF. The binding site in Switch I remains flexible, which enables binding, albeit with ∼10-fold lower affinity, and remarkably, this interaction with BRAF reverses the compound’s blockage of intrinsic GTP hydrolysis. Unlike KRAS^WT, GDP-loaded KRAS^G12D surprisingly maintains a low-affinity interaction with BRAF-RBD, but MRTX1133 can circumvent this mutant-specific abnormal interaction. Taken together, MRTX1133 allosterically ‘freezes’ the KRAS^G12D nucleotide-binding site conformation, arresting the canonical GTPase cycle of this oncogenic mutant. This provides a framework for understanding the mechanisms-of-action of SII-P-directed inhibitors and how tumours may acquire resistance.</div></div>","PeriodicalId":369,"journal":{"name":"Journal of Molecular Biology","volume":"437 14","pages":"Article 169162"},"PeriodicalIF":4.7,"publicationDate":"2025-04-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143908289","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}

{"title":"BEAGLE 2.0: A Web Server for RNA Secondary Structure Similarity Detection Leveraging SHAPE-directed RNA Structure Determination","authors":"F. Ballesio ,&nbsp;A. Teofani ,&nbsp;C. Carrino ,&nbsp;M. Catalano ,&nbsp;M.L. Nicolaeasa ,&nbsp;G. Ausiello ,&nbsp;M. Helmer-Citterich ,&nbsp;P.F. Gherardini","doi":"10.1016/j.jmb.2025.169154","DOIUrl":"10.1016/j.jmb.2025.169154","url":null,"abstract":"<div><div>Recent studies underscore the significant role of RNA secondary structures in various biological and pathological processes. Structural conservation can reveal homologies undetectable by sequence analysis alone, making accurate prediction and comparison of RNA secondary structures crucial. The BEAGLE algorithm enables pairwise alignments of RNA secondary structures through dynamic programming, leveraging the BEAR encoding for RNA secondary structures representation. Initially, BEAGLE was designed to perform pairwise alignments of user-provided RNAs or against a limited number of datasets. We now introduce BEAGLE 2.0, a web server designed to facilitate the search for structural similarities between user-provided RNA molecules and an expanded collection of RNA secondary structure datasets. These datasets include structures derived from SHAPE experiments in <em>Homo sapiens</em>, <em>Mus musculus</em>, <em>Danio rerio</em>, <em>Escherichia coli</em>, <em>Bacillus subtilis</em>, and various viruses, including SARS-CoV-2. It also incorporates predicted structures from the NONCODE database for a wide range of animals and plants, as well as a dataset of structures based on constraints derived from conserved positions within the families present in Rfam. Users can input RNA sequences or a combination of sequences and secondary structures in either dot-bracket or BEAR format. BEAGLE 2.0 outputs pairwise alignments with measures of structural similarity and statistical significance. Additionally, it offers a visual representation of the secondary structures, with structural elements highlighted in different colors. Overall, BEAGLE 2.0 enables searches in RNA structure datasets leveraging experimentally supported data, to identify structural similarities in RNAs of interest. BEAGLE 2.0 is available at <span><span>https://beagle2.bio.uniroma2.it</span><svg><path></path></svg></span>.</div></div>","PeriodicalId":369,"journal":{"name":"Journal of Molecular Biology","volume":"437 15","pages":"Article 169154"},"PeriodicalIF":4.7,"publicationDate":"2025-04-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143952132","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}

{"title":"DINC-ensemble: A web server for docking large ligands incrementally to an ensemble of receptor conformations","authors":"Anja Conev ,&nbsp;Jing Chen ,&nbsp;Lydia E. Kavraki","doi":"10.1016/j.jmb.2025.169163","DOIUrl":"10.1016/j.jmb.2025.169163","url":null,"abstract":"<div><div>Protein–ligand docking aids structure-based drug discovery by computationally modelling protein–ligand interactions. DINC (Docking INCrementally) is one approach to molecular docking that improved the docking of large ligands using a parallelized incremental meta-docking. Traditional docking tools, including DINC, explore the flexibility of the ligand in a single receptor binding pocket assuming limited flexibility of the receptor backbone. This simplifying assumption narrows down the docking search space but hinders successful docking for flexible receptors. DINC-Ensemble implicitly considers receptor backbone flexibility by running DINC docking in parallel on different receptor conformations. Inputs to DINC-Ensemble include (1) a ligand and (2) a list of different receptor conformations. For each ligand-receptor pair DINC-Ensemble performs incremental meta-docking in parallel. As a result, multiple ligand poses are generated in the binding pockets of different receptor conformations. These poses are then ranked, and the lowest scoring pose is selected. Two main outputs provided by a successful run of DINC-Ensemble are (1) the best scoring ligand poses and (2) a ranked list of selected receptor conformations. The best scoring ligand pose can be used to understand the interactions between the receptor and the ligand that influence the binding. The ranked list of receptor conformations shows the best receptor conformation fit for a given ligand and can provide insight into ligand-induced conformational selection. We provide DINC-Ensemble as a Python package and a free web server at <span><span>https://dinc-ensemble.kavrakilab.rice.edu/</span><svg><path></path></svg></span>.</div></div>","PeriodicalId":369,"journal":{"name":"Journal of Molecular Biology","volume":"437 15","pages":"Article 169163"},"PeriodicalIF":4.7,"publicationDate":"2025-04-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144053468","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}