Results
- Showing results for:
- Reset all filters
Search results
-
Journal articleWan Y, Wong JLC, Sanchez-Garrido J, et al., 2026,
Genomic and molecular characterisation of a KPC-producing Klebsiella pneumoniae clinical isolate resistant to meropenem-vaborbactam, imipenem-relebactam, and ceftazidime-avibactam
, BMC Genomic Data, Vol: 27, ISSN: 2730-6844Background Resistance to carbapenems and third-generation cephalosporins is increasing in Klebsiella pneumoniae globally, restricting therapeutic options. The β-lactam/β-lactamase inhibitor combinations are widely used to circumvent β-lactamase-mediated resistance. In 2021, an unusual K. pneumoniae clinical isolate, KpMVR1, was recovered from a hospitalised patient in England, exhibiting resistance to meropenem-vaborbactam, imipenem-relebactam, and ceftazidime-avibactam. To investigate this phenomenon, we characterised the genome and antimicrobial susceptibility of KpMVR1 alongside two clonally related isolates susceptible to all three β-lactam/β-lactamase inhibitor combinations: KpMVS1, collected from the same patient 42 days earlier, and KpMVS2, from another patient in the same hospital. Methods Illumina and MinION whole-genome sequencing were conducted for these three isolates, followed by hybrid genome assembly. Annotated genome assemblies were compared to identify genetic variation. Mutagenesis experiments were performed to verify predicted functional alterations. Results All isolates belonged to clone ST8134 and carried blaKPC-2 alleles (KpMVR1: blaKPC-157; KpMVS1 and KpMVS2: blaKPC-2) in plasmids predicted to be conjugative. Insertion sequence ISEc68 caused a frameshift mutation in KpMVR1’s ompK36 gene, reducing susceptibility to meropenem-vaborbactam and imipenem-relebactam. KPC-157 demonstrated decreased hydrolysis of imipenem and ceftazidime when compared with KPC-2. KpMVR1 also encoded a disrupted transcriptional repressor MarR and a destabilising mutation in AcrB, a component of the AcrAB-TolC multidrug efflux pump. An intact, iron-transporting fec operon was identified on a novel IncFII(pKP91)/IncFIB(K) plasmid unique to KpMVS2, possibly accounting for the cefiderocol resistance observed in this isolate. ConclusionsKpMVR1 carried multiple resistance-associated genetic alterations and likely developed its resistance profile
-
Journal articleYi L, Kulik N, Shao S, et al., 2026,
The transmembrane domain is a primary determinant of the location and substrate specificity in cyanobacterial FtsH heterocomplexes
, Algal Research, Vol: 98, ISSN: 2211-9264Membrane-embedded FtsH proteases play important and diverse physiological roles in prokaryotes, chloroplasts and mitochondria, but how substrates are distinguished remains unclear. The cyanobacterium Synechocystis sp. PCC 6803 contains four FtsH homologs organized into two distinct heterocomplexes and one homocomplex. The two heterocomplexes, derived from a recent gene duplication event, are the essential FtsH1/3 complex found in the cytoplasmic membrane, and the thylakoid-embedded FtsH2/3 complex, with a role in the repair of photodamaged photosystem II. Using a domain swapping approach, we demonstrate here that the transmembrane domains of FtsH1 and FtsH2 are primary determinants of the cellular location and functional differences between the FtsH1/3 and FtsH2/3 complexes, whereas the soluble AAA+ (ATPases associated with diverse cellular activities) and protease domains and the soluble linker were largely interchangeable under the conditions tested. Overall, our findings identify the transmembrane domain as an important determinant of both the location and substrate specificity of FtsH heterocomplexes and support a role for this domain in their functional evolution.
-
Journal articleSchröder GC, Crichlow GV, Jablonowski E, et al., 2026,
Identification of a key water molecule involved in the macrophage migration inhibitory factor‐catalyzed tautomerization of para‐hydroxyphenylpyruvate using neutron crystallography
, Protein Science, Vol: 35, ISSN: 0961-8368Neutron crystallography was used to determine a 2.5-Å resolution all-atom structure of macrophage migration inhibitory factor (MIF) interacting with 3-(4-hydroxyphenyl)-pyruvate (HPP). MIF is a pro-inflammatory, pro-tumorigenic protein that may be an attractive therapeutic target. MIF catalyzes the interconversion of the keto and enol forms of HPP by a tautomerase reaction. Although HPP is evidently not a physiological substrate of MIF, many compounds that inhibit this activity in enzymatic assays have been found also to inhibit physiological activities of MIF. Therefore, the MIF-catalyzed HPP tautomerization reaction is used in initial screening of compounds in the search for inhibitors of MIF physiological activity. The neutron diffraction-derived crystal structure reveals the position of a water molecule involved in the tautomerization reaction, and also confirms the charged state of lysine-32 in the active site. The structure confirms the previously proposed catalytic mechanism of MIF, with the N-terminal Pro-1 abstracting a proton to generate an HPP enolate intermediate which is subsequently protonated. The structure reported herein reveals that this proton is supplied by a neighboring water molecule. Along with the neutron structure, a room-temperature synchrotron x-ray crystal structure reveals a covalent adduct between HPP and MIF. While this adduct is a result of radiation-induced chemistry, its formation confirms the catalytic role of the active site residue because a covalent complex could only form if the reactive carbon of the substrate is correctly positioned by the enzyme.
-
Journal articleFeng N, Zhang X, Chen K, et al., 2026,
Generalized z-Domain Modeling and Analysis for High-Order Grid-Connected Inverters
, IEEE Transactions on Power Electronics, Vol: 41, Pages: 15649-15664, ISSN: 0885-8993With the rapid development of power electronics technology, discrete-time digital controllers are taking an increasingly dominant role in converter control. Existing converter control methods lack the scalability required by practical power electronic systems with high-order filters and lack the accuracy needed for stability assessment under complex topologies. To remove these limitations, this article proposes a highly accurate and scalable method for linear time-invariant systems of arbitrary integer order that include time delays. The proposed approach utilizes the so-called modified z-transform that accurately models delays that are not necessarily multiples of the sampling period, and therefore yields an enhanced capability for stability margin assessment. The z-domain modeling, digital controller design, and stability analysis are discussed in detail and compared with mainstream methods. The improved performance of the proposed method is verified via both numerical simulations and physical experiments, under various component configurations and filter topologies. The results show that the proposed method predicts the margin of the stabilizing feedback gain significantly more accurately than the ones predicted by the existing methods.
-
Journal articleMurray JW, 2026,
Random repeats open a route into folded protein space.
, Proc Natl Acad Sci U S A, Vol: 123 -
Journal articleFrankel G, 2026,
Design of a live-attenuated bacterial vaccine using effector network engineering
, Nature Communications, ISSN: 2041-1723Enteropathogenic Escherichia coli (EPEC) and Citrobacter rodentium (CR) are extracellular enteric attaching and effacing (A/E) pathogens of humans and mice, respectively. Their virulence relies on intimate bacterial attachment and a network of type III secretion system effectors. Here, through systematic reduction and redesign of the effector network in CR, we develop an attenuated strain, CRV (CR Vaccine), encoding a subset of ten effectors. CRV colonises C57BL/6 mice ~100-fold lower than wild type CR (CRWT) without causing overt pathogenesis. Moreover, C3H/HeN mice, which succumb to CRWT infection, survive CRV challenge. Vaccination with CRV confers protection against subsequent CRWT infection in both mouse strains. Serological analysis reveals a repertoire of dominant CR antigens, including the O-antigen, the virulence factors intimin, EspA and Tir and the outer membrane proteins Lpp, OmpA, MetQ and CARC (an AIDA-like autotransporter). We show that CRWT and CRV immunisation elicits comparable B cell and antibody responses, which is contingent on intimate bacterial attachment. A corresponding EPEC strain (E. coli Vaccine, ECV) effectively colonises epithelial cells in a gut-on-chip model. These findings establish effector network minimisation as a generalisable strategy for rational bacterial attenuation and live-attenuated vaccine design.
-
Journal articleWong LH, Mitchell NA, Heng JYY, 2026,
Population balance modelling case studies for protein crystallisation: Workflows addressing impurities and heterogeneous seeding
, Chemical Engineering Research and Design, Vol: 232, Pages: 544-565, ISSN: 0263-8762In this study, a population balance modelling (PBM) workflow for protein crystallisation is introduced and applied to two case studies, modelling lysozyme crystallisation in the presence of protein impurities (thaumatin and bovine serum albumin (BSA)) and heterogeneous silica seeds. The workflow systematically applies engineering judgement to identify phenomena (nucleation, growth, and enhancement/inhibition by foreign species), incorporates linear and exponential empirical adjustments to traditional kinetic models, and employs sequential kinetic parameter estimation to quantify phenomena. Across the two studies, workflow applicability was demonstrated for different crystalliser configurations (static/orbital shaking), buffer systems, foreign species, and scale-up (1–5 mL), whilst also overcoming experimental and solubility data limitations often encountered in high-cost crystallisation experiments. Although some calibrated parameters were uncertain with the 95% t-value smaller than the reference (attributed to a need for larger datasets), the workflow not only increased system knowledge but also revealed a critical knowledge gap: the complex, protein impurity-dependent behaviour of ternary lysozyme-silica-impurity systems, which traditional kinetic models proved unable to capture. The proposed workflow is broadly applicable to other peptides, proteins, or even small molecules, positioning it as a potential method for accelerating process design and understanding.
-
Journal articleManchinu MF, Congiu M, Massidda M, et al., 2026,
PBMC DEG/miRNA biomarkers of TDP-43 pathology in ALS.
, Neurobiol Dis, Vol: 226Amyotrophic lateral sclerosis (ALS) lacks reliable, disease-specific, and minimally invasive biomarkers, representing a major barrier to early diagnosis and patient stratification. The primary aim of this translational pilot study was to identify a disease-specific, TDP-43-related, gene-microRNA (miRNA) signature in peripheral blood mononuclear cells (PBMCs) of ALS patients with potential diagnostic value. To this end, we first identified differentially expressed disease-specific genes (dsDEGs) using a TDP-43-based rat model of ALS, generated by stereotaxic infusion of full-length (FL) TAR DNA-binding protein 43 (TDP-43) into the motor cortex. Transcriptomic profiling of the motor cortex revealed candidate dsDEGs, which were subsequently validated by RT-qPCR in motor cortex, spinal cord, and PBMCs from the same animals. To assess translational relevance, expression levels of these dsDEGs were analyzed in PBMCs from early- to mid-stage ALS patients and matched healthy controls, while disease specificity was evaluated using Parkinson's disease (PD) samples. In parallel, conserved miRNAs predicted to target the identified dsDEGs were examined in both rat and human PBMCs. Five dsDEGs, Mctp1, Penk, Mt2A, Drd1, and Rasgrp2, were consistently dysregulated across central and peripheral tissues in the TDP-43 rat model. RT-qPCR analysis of human PBMCs confirmed significant and selective dysregulation of these genes in ALS, but not in PD, supporting disease specificity. Moreover, exposure of human neuroblastoma cells and healthy PBMCs to TDP-43 recapitulated the ALS-like expression changes. Computational and experimental analyses identified seven conserved miRNAs targeting these dsDEGs, of which four were significantly downregulated in ALS PBMCs, supporting a coordinated regulatory network. Receiver operating characteristic (ROC) analyses demonstrated strong discriminative performance for both the gene signature (AUC 0.87-1.00) and the associated miRNAs (AUC 0.95-1.00). Togeth
-
Journal articleSpeck C, Reuter M, Weekes C, 2026,
Mechanisms of MCM2-7 helicase activation and initial DNA melting at near base-pair resolution
, Nature Communications, ISSN: 2041-1723During eukaryotic DNA replication initiation, inactive MCM2–7 double-hexamers assembled at replication origins must be converted into two active CMG helicases, yet how this transition is coupled to origin DNA unwinding in vivo remains unclear. Here, we identify a DNA-bound intermediate with an extended genomic footprint that forms during helicase activation. Genome-wide mapping of initial strand separation reveals that DNA unwinding initiates near the N-terminal interface of opposing MCM2–7 hexamers. At these sites, the origin DNA exhibits a conserved AT-rich/GC-rich/AT-rich sequence architecture centred under the helicase complex, which is associated with an elevated DNA melting probability. We further show that restricting hexamer splitting delays release of the Cdc45-loading factor Sld3, demonstrating that mechanical transitions during helicase activation are tightly coupled to complex disassembly. Finally, we provide in vivo evidence that single-stranded DNA is ejected through a specialised DNA exit gate at the Mcm2/5 interface during helicase activation, which is dispensable for ongoing DNA synthesis. Together, these findings establish a mechanistic framework for how replication origins are remodelled to initiate DNA replication and reveal key intermediates and DNA transactions during helicase activation.
-
Journal articleHobbs B, Limmer N, Clenshaw GL, et al., 2026,
Accurate interdomain contacts in a mixed folded protein from NMR-guided coarse-grained simulations.
, Phys Chem Chem Phys, Vol: 28, Pages: 17194-17202Intrinsically disordered, low-complexity regions frequently cooperate with folded domains to mediate protein-protein interactions, yet accurately describing these mixed folded-disordered systems remains challenging. To visualize these mixed folded proteins, experimentally guided coarse-grained (CG) molecular dynamics simulations are often employed to extend the timescales required to capture the complex dynamics in play. However, the minimalistic nature of these approaches often compromises structural accuracy and can lead to inaccurate inter-domain interactions. Here we introduce backbone dihedral terms directly derived from NMR chemical shift data in CG-simulations to characterize the open state of a mixed-folded construct of the anti-aggregation chaperone DNAJB6 that contains a folded J-domain and a disordered GF linker. By tuning residue-specific backbone dihedral parameters to match NMR-derived secondary-structure propensities of the linker in CG-simulations, we generate conformational ensembles that yield accurate interdomain contact maps. In agreement with analysis of NMR relaxation data, the resulting ensembles show that even in the nominally open state the linker experiences motions that resemble those of the closed state driven by hydrophobic residues in GF. More generally, we show that by expanding CG-simulations to allow them to capture both local and global structural properties, physically relevant interdomain contacts can be retrieved.
-
Journal articleNoguchi Y, Speck C, Saleh A, 2026,
Structural insights into Sld3-Sld7-dependent Cdc45 loading during replication initiation
, Nature Communications, ISSN: 2041-1723How DDK phosphorylation primes the MCM2-7 double hexamer (DH) for Sld3-Sld7 binding and Cdc45 loading during helicase activation remained unclear. We define this mechanism through cryo-EM structures of MCM2-7 DH-Sld3-Sld7 (MS) and MCM2-7 DH-Sld3-Sld7-Cdc45 (MSC). Our reinterpretation of published DH maps reveals that the autoinhibitory Mcm4 tail engages sites on both Mcm4 and Mcm6, extending the known autoinhibitory region. Upon DDK-dependent phosphorylation, both sites become accessible. In the context of the MS structure, we discovered that two short Sld3 motifs (MCM recognition domain MRD1 and MRD2) contact now Mcm4 and Mcm6, to read out DH phosphorylation state, while Sld7 anchors the DH at the Mcm2/Mcm6 interface via an MCM-binding helix, and the Sld3 Treslin domain (STD) binds Mcm2. We show that Sld3-Sld7 forms a heterotetramer in solution, but in MS, the Sld7 dimerization domains become separated. In the MSC structure, Cdc45 dislodges the Sld3 STD from Mcm2, allowing Sld3 to position Cdc45 at the Mcm2/Mcm5 interface while MRD1/2 retain their Mcm4/Mcm6 contacts, rationalizing Sld3-dependent Cdc45 recruitment. Mutagenesis of the Sld3 STD-Cdc45 interface disrupts Cdc45 loading, validating this interaction. Together, our data reveal a phosphorylation-encoded mechanism coupling DDK-activated Mcm4/Mcm6 surfaces to distal Cdc45 placement, explaining how firing factors choreograph the DH-to-CMG transition.
-
Journal articleSong G, Ma Z, Fan M, et al., 2026,
Internalized components of membrane attack complexes disrupt proteostasis and acquire alarmin-like properties.
, Nat CommunImmune effects of membrane attack complexes (MAC) have been widely attributed to their abilities to cause cell death. Here, we show that the MAC component, C9, forms non-cytolytic aggregates with pro-inflammatory effects. Intracellular aggregates of C9 are detected within inflamed tissues of patients in association with endothelial cell (EC) activation but not increased cell death. We identify NUMBL as a Rab35 effector that directly binds surface-bound C9 to promote C9 internalization and entry into the endolysosomal pathway. Within acidified endolysosomes, C9 forms insoluble aggregates that are targeted for degradative aggrephagy in a process that activates NF-κB. For C9 aggrephagy to occur, ZFYVE21, a Rab5 effector, complexes with RNF34 to bridge C9 aggregates to LC3B+ aggresome membranes. We detect C9 aggregates in vivo, and we show that a ZFYVE21-RNF34 signaling axis is required for C9 aggrephagy and NF-κB -dependent EC activation in three separate mouse models. Mice with conditional loss of ZFYVE21 in ECs show reduced aggregraphy, resulting in attenuated systemic inflammation and reduced tissue injury following skin transplantation. Our data show that the C9 component of MACs forms intracellular aggregates with alarmin-like properties.
-
Journal articleLau R, Giblin S, Sugar A, et al., 2026,
SpyCEP dismantles neutrophil immunity via disorder-driven chemokine remodeling and GAG targeting
, Proceedings of the National Academy of Sciences of the United States of America, Vol: 123, ISSN: 0027-8424Streptococcus pyogenes evades neutrophil-mediated immunity by secreting the protease SpyCEP, which inactivates chemokines such as CXCL8; however, the mechanism by which SpyCEP targets CXCL8 for cleavage has remained unclear. This work uncovers an intrinsically disordered autocatalytic maturation loop that binds CXCL8 and induces a conformationally heterogeneous state in the chemokine. A model is proposed in which this disorder-mediated recognition facilitates access to the substrate cleavage site and is compatible with SpyCEP acting at glycosaminoglycan (GAG)-bound CXCL8 reservoirs. This disorder-mediated mode of substrate recognition departs from classical protease–substrate interfaces and identifies the SpyCEP cleaved autocatalytic matu-ration loop (CAML) as a potential target for anti-virulence strategies against S. pyogenes.
-
Journal articleZhao Z, Vercellino I, Whitelegge JP, et al., 2026,
Cryo-EM structures of naturally occurring dimeric photosystem II complexes lacking the Mn₄CaO₅ cluster
, Nature Communications, ISSN: 2041-1723Robust oxygenic photosynthesis requires the efficient assembly and repair of the multi-subunit oxygen-evolving photosystem II (PSII) complex. Previous cryogenic electron microscopy (cryo-EM) structures of PSII assembly/disassembly intermediates have relied on the analysis of deletion mutants or removal of PSII subunits in vitro. Here we report the cryo-EM structures of naturally occurring dimeric PSII intermediates from the cyanobacterium Thermosynechococcus vestitus at a resolution of about 2.2 Å. These intermediates contain inactive dimers lacking the oxygen-evolving complex (OEC) and semi-active dimers with the OEC present in one of the two monomers. Our structural data provide a mechanism for how assembly and disassembly of the Mn4CaO5 cluster is coordinated with the binding and release of the extrinsic proteins: restructuring of the C-terminal tail of D1 subunit during assembly or disassembly of the Mn cluster triggers conformational changes in D2, CP47 and CP43 to drive the binding/release of the extrinsic proteins. A combination of structural and mass spectrometry data also suggests that the inactive PSII complexes may include damaged complexes containing oxidized D1-His332, a monodentate ligand to one of the Mn ions of the OEC.
-
Journal articleLi X, Xu L, He F, et al., 2026,
Precise Control of Photomechanical Response Modes of Barbituric Acid Derivative Crystals: from Bending to Fragmentation
, Crystal Growth and Design, Vol: 26, Pages: 5242-5251, ISSN: 1528-7483Photomechanically responsive crystals show significant application potential in fields such as flexible optoelectronic devices, micro/nano manipulation, and smart biomimetic materials. However, how to precisely control the photomechanical response modes of crystals (such as bending and fracturing) through molecular design, remains a major challenge in this field. This paper reports four novel barbituric acid derivative crystals (BTBA, DBH-DMB, DCMP, and DCB-TP), which exhibit distinct photoinduced bending and photoinduced fracturing behaviors upon light excitation. Calculations of single-crystal structures, Hirshfeld surfaces, and energy frameworks indicate that BTBA and DBH-DMB, which have smaller molecular dihedral angles, possess two-dimensional layered sliding structures formed by strong π–π stacking. The internal stress generated by photoexcitation can be dissipated through the ordered sliding of molecular layers, thereby exhibiting photoinduced bending behavior; in contrast, DCMP and DCB-TP, which have nearly perpendicular molecular dihedral angles, exhibit larger steric hindrance that impedes π–π stacking and lacks sliding interfaces; internal stress accumulates rapidly at defects, ultimately leading to crystal fragmentation. Notably, the bending behavior of BTBA is partially thermally reversible, whereas that of DBH-DMB is irreversible. Size effects indicate that longer and thinner crystals exhibit lower bending stiffness and superior tip deflection performance. Molecular orbital energy levels further reveal that bending crystals possess moderate energy gaps and significant intramolecular charge transfer characteristics, which are conducive to the formation of oriented stress gradients; in contrast, fracturing crystals have larger energy gaps, higher degrees of electronic localization, and greater structural rigidity. This study uncovers the synergistic regulatory mechanisms by which molecular conformation, stacking patterns, and elec
-
Journal articleGottweis J, Weng W-H, Daryin A, et al., 2026,
Accelerating scientific discovery with Co-Scientist
, Nature, ISSN: 0028-0836Scientific discovery is driven by scientists generating hypotheses for complex problems that undergo rigorous experimental validation. To augment this process, we introduce Co-Scientist, a multi-agent artificial intelligence (AI) system built on Gemini for structured scientific thinking and hypothesis generation. Co-Scientist aims to help scientists discover new original knowledge. Conditioned on their research objectives and previous scientific evidence, it formulates demonstrably novel research hypotheses for experimental verification. The system’s design involves agents continuously generating, critiquing and refining hypotheses accelerated by scaling test-time compute. Key contributions include (1) a multi-agent architecture with an asynchronous task execution framework for flexible compute scaling, and (2) a tournament evolution process for self-improving hypotheses generation. Automated evaluations show continued benefits of test-time compute scaling, improving hypothesis quality over time. Although this is a general-purpose system, we focus the validation in three biomedical applications: drug repurposing; novel-target discovery1; and explaining mechanisms of antimicrobial resistance2. Specifically, Co-Scientist helped to identify new drug-repurposing candidates and synergistic combination therapies for acute myeloid leukaemia that were validated through in vitro experiments. These real-world validations demonstrate the potential of Co-Scientist to accelerate scientific discovery and usher in an era of AI-empowered scientists.
-
Journal articleParacuellos P, Bexter A, Patkowski JB, et al., 2026,
Molecular basis of type VI secretion system effector loading
, Nature Microbiology, Vol: 11, Pages: 1992-1994, ISSN: 2058-5276Type VI secretion systems (T6SSs) are widespread bacterial nanomachines that deliver effectors into prokaryotic and eukaryotic cells. How an effector cargo is recruited and loaded into the Hcp ring assemblies that form the tube injected by the T6SS remains poorly understood. Pseudomonas aeruginosa has four T6SSs, each associated with a different Hcp protein. Here we use cryo-electron microscopy to resolve the structure of the Tce1 cargo loaded into a Hcp3 ring from the P. aeruginosa H3-T6SS. We show that a single Tce1 monomer interacts asymmetrically with, and is enclosed by, two hexameric Hcp3 rings, engaging key residues lining the inner surface of the Hcp3 disc. Our data indicate a stepwise loading mechanism, where an initial heterodimeric Hcp–cargo complex forms before ring encapsulation around the effector. Structural modelling suggests similar effector–Hcp3 interactions for a second T6SS effector, Tce2, which has antifungal activity. We propose that this mechanism enables coordinated delivery of a broad payload into target cells.
-
Journal articleCosta TRD, Penadés JR, 2026,
Escaping our own biases: AI, scientific reasoning, and discovery in molecular biology
, Molecular Cell, ISSN: 1097-2765Artificial intelligence may transform molecular biology not by always providing the correct answer, but by exploring mechanistic possibilities unconstrained by the assumptions that shape expert thinking. Its greatest contribution may be exposing the intellectual blind spots of scientists, while experimental validation remains the ultimate arbiter of truth.
-
Journal articleArino S, Fusco G, De Simone A, 2026,
α-Synuclein aggregation landscape from phase separation to neurotoxic intermediates.
, FEBS Lett, Vol: 600, Pages: 1955-1965The aberrant aggregation of α-synuclein (αS) into insoluble amyloid fibrils is a hallmark of Parkinson's disease. Despite recent advances in characterising the properties of mature αS fibrils, the transient and heterogeneous intermediates that underlie cellular toxicity remain largely elusive. Here, we review the mechanistic principles of αS aggregation, focussing on liquid-liquid phase separation (LLPS) as a critical intermediate step. We discuss how the structural evolution of αS within the condensed phase governs the subsequent patterns of cellular dysfunction and pathological propagation. This framework supports an emerging state-centric paradigm in therapeutic discovery, where the physical properties of αS condensates are modulated to mitigate the deleterious effects of its misfolding, offering a new sophisticated alternative to classical inhibition strategies.
-
Journal articleSanchez-Garrido J, David S, Rattle J, et al., 2026,
Type 3 fimbrial regulation underpins anti-MrkA immunotherapeutic efficacy in experimental Klebsiella pneumoniae infection.
, J Infect DisBACKGROUND: Klebsiella pneumoniae (KP) is a critical-priority organism due to prevalent last-line antibiotic resistance. Alternative treatments, including vaccines and monoclonal antibodies (mAb), depend on antigen (Ag) expression at infection sites for immunotherapeutic activity. However, the relationship between genome-encoded Ag presence and Ag expression is often overlooked. Here, we use the KP type 3 fimbrial (T3F) subunit MrkA as a prototype to build a generalisable framework to assess Ag expression and its correlation with in vivo immunotherapeutic efficacy. METHODS: We perform genomic analysis of 1649 KP genomes for T3F genes, including structural and regulatory components. We generate isogenic mutants with absent, normal or overexpressed MrkA and profile MrkA expression at single-cell level from murine pneumonia and bacteraemia models. We compare anti-MrkA mAb efficacy in vivo against strains with normal and enhanced MrkA expression. RESULTS: T3F structural and regulatory genes are highly conserved, however, regulatory gene disruption (mrkH) is more common than structural gene disruption and, in both cases, MrkA Ag is not expressed. In vivo Ag profiling revealed site-specific differences in MrkA expression, with ∼20% of KP cells expressing MrkA in the lung versus ∼5% in the bloodstream. Anti-MrkA mAb activity was dependent on MrkA abundance, with significantly enhanced efficacy following infection with MrkA-overexpressing KP. CONCLUSIONS: Regulatory genes are as important to characterise as structural gene presence when evaluating antigen candidates in clinical isolates, and Ag expression can vary by anatomical context. For MrkA, Ag abundance determines anti-MrkA mAb activity, suggesting infections with high MrkA expression will respond better to therapy.
-
Journal articleEttema TW, Inaba-Inoue S, Thangaratnarajah C, et al., 2026,
Shared structural mechanisms of alternating access between the secondary peptide transporter SbmA and ABC transporters
, Nature Communications, Vol: 17, ISSN: 2041-1723SbmA is a membrane transporter from Escherichia coli that imports antimicrobial peptides. SbmA belongs to the SbmA-like peptide transporter (SLiPT) family. Although the protein is a secondary active transporter that is energized by the proton gradient, it is structurally related to the transmembrane domain (TMD) of ATP-binding cassette (ABC) transporters. SbmA therefore bridges the structural divide between primary and 61 secondary transporters. However, it remains unclear, if SbmA also shares the mechanism of alternating access with ABC transporters, because only a single (outward-open) state is resolved. Here, we show by sequence analysis that SbmA is likely evolved from the TMD of an early ancestor of the ABC transporter YddA. We determine the cryogenic electron microscopy structures of SbmA in occluded and inward-facing states. These conformations closely resemble equivalent states found in ABC trans- porters, indicating a shared structural mechanism of transport. In contrast to ABC transporters, where nucleotide binding, hydrolysis and release steer conformational changes necessary for substrate translocation, electron paramagnetic resonance (EPR) spectroscopy and molecular dynamics (MD) simulations reveal how pH changes induce conformational transitions in SbmA, consistent with a mechanism of substrate internalization that utilizes the transmembrane proton gradient.
-
Journal articleGroff A, Lu Y, Feeney M, et al., 2026,
Sustainable production of myoglobin meat protein in plant chloroplasts
, Frontiers in Plant Science, Vol: 17, Pages: 1-15, ISSN: 1664-462XAlternative routes for producing animal proteins are crucial for reducing the reliance on traditional livestock farming, which contributes significantly to greenhouse gas emissions, deforestation, and water consumption. Myoglobin (Mb) is an important oxygen-binding hemoprotein found in vertebrate muscle, that enhances the nutritional and sensorial properties of meat. Due to its unique functionality, Mb has been heterologously expressed in a variety of organisms, although only transient expression in Nicotiana benthamiana has been reported for higher plants. In this study, we used chloroplast transformation technology to express porcine Mb in higher plants (tobacco, a non-edible model plant, and lettuce, an edible host) and bovine Mb in the green alga Chlamydomonas reinhardtii. Mb accumulation was estimated by immunoblotting and found to be much higher in tobacco (2.7% total soluble protein (TSP)) and lettuce (1.5% TSP) than Chlamydomonas reinhardtii (<0.25% TSP). The expression in tobacco chloroplasts is also superior to tobacco nuclear expression (using either the cauliflower mosaic virus 35S promoter or Arabidopsis thaliana ubiquitin promoter). Total heme levels were elevated in myoglobin-producing mutants compared to control plants, although porcine Mb purified from tobacco leaves exhibited approximately 35% heme-binding (compared to 80% heme-binding in E. coli-expressed Mb), despite being correctly folded, suggesting that heme availability might be a bottleneck. Overall, our work describes the first report of stable Mb production in higher plants and its effect on photosynthesis and heme levels. This provides a foundation for future plant-made animal proteins for food applications.
-
Conference paperPessina D, Heng JYY, Papathanasiou MM, 2026,
An in silico/in vitro approach for uncertainty-aware hybrid models for template-induced protein crystallisation systems
, The 36th European Symposium on Computer Aided Process Engineering, Publisher: PSE Press, Pages: 631-639, ISSN: 2818-4734<jats:p>Crystallisation is a promising and scalable alternative to chromatography for biologics purification. However biologics such as proteins and peptides often crystallise only in narrow operating windows, limiting process flexibility. Template-induced crystallisation can lower supersaturation requirements and expand feasible operating ranges, yet the template dependence of nucleation and growth kinetics remains difficult to parametrise mechanistically. To address this, we develop and experimentally validate uncertainty-aware hybrid models for lysozyme crystallisation on hydroxyl- and carboxyl-functionalised silica templates. A mechanistic population-balance model is coupled to a data-driven regressor that maps operating conditions and template variables to effective nucleation and growth rates. We compare a neural network baseline against a structured neural power-law surrogate, which embeds a supersaturation-dependent power-law form. Both hybrid models are trained in-the-loop via differentiable simulation, and variational inference is used to obtain posterior parameter distributions and calibrated predictive uncertainty. Across cross-validation and off-grid tests at previously unseen combinations of temperature and template loading, the hybrid models accurately reproduce solute concentration dynamics and capture key particle-size trends, while the neural power-law surrogate provides improved robustness and faster uncertainty quantification. These results support hybrid, uncertainty-aware PBMs as practical tools for prediction, design-space exploration, and comparison of template-enabled protein crystallisation processes.</jats:p>
-
Journal articleFrankel G, 2026,
Phospholipid-1 independent biogenesis and function of the RP4 conjugation pilus
, Nature Communications, ISSN: 2041-1723Bacterial conjugation, the process of horizontal gene transfer between bacteria, is initiated by mating pair formation (MPF) via a conjugative pilus. Conjugation of the IncP RP4 plasmid is mediated by short mating pili. Here, we report the cryo-EM structure of the RP4 pilus at 2.74 Å resolution. Uniquely, both the structural and quantitative mass spectral analyses revealed that the cyclic TrbC pilin subunit is not lipidated. Consistently, an E. coli pgsA mutant lacking phosphatidylglycerol (PG) can serve as a donor of RP4 but not of F- (pKpQIL), H- (R27) or W- (R388) pili, whose biogenesis and DNA transfer is PG-dependent. RP4 is the first example of a lipid-independent functional mating pilus. This discovery suggests that an amphipathic lipid moiety is not universally essential for the biogenesis of conjugative pili and MPF, providing an alternative model for their assembly and function. These data expand our understanding of the diverse bacterial mechanisms employ to transfer genetic material.
-
Journal articleRibardo DA, Singh NK, Beeby M, et al., 2026,
The Campylobacter jejuni flagellar V-ring discerns viscosity levels to alter swimming velocity, metabolic gene expression, and fitness.
, Proc Natl Acad Sci U S A, Vol: 123Campylobacter jejuni is an intestinal commensal of birds and animals and a leading cause of bacterial diarrheal disease in humans. In hosts, C. jejuni primarily resides in the mucus layer atop the lower intestinal epithelium. Persistence in this niche requires a single flagellar motor at both C. jejuni poles that generates high torque for flagellar rotation to facilitate motility and high swimming velocities. Unlike many bacterial flagellates, C. jejuni swimming velocity increases as external viscosity increases. We identified a complex formed by FlgV, VidA, and VidC (Cjj81176_1732) positioned near the MS-ring-rotor junction in the flagellar motor we annotated as the V-ring. Viscosity-influenced growth, modulation of swimming velocity, and transcription of iron/heme acquisition, respiratory, and energy-generating systems were dependent on the V-ring. C. jejuni ΔflgV and ΔvidC populations lacking a complete V-ring were motile, but could not optimally modulate swimming velocity. Like nonmotile flagellar stator or filament mutants, motile V-ring mutants had in vivo and in vitro growth and viability defects and dysregulated transcription of genes likely impacting physiology. Because the V-ring mutants behaved similarly as nonmotile mutants that experience little to no viscous drag on the filament, we propose C. jejuni V-ring mutants cannot detect viscous drag on their rotating filaments. We propose the V-ring evolved in C. jejuni and potentially other bacteria producing high torque flagellar motors to monitor external viscosity information via viscous drag on the rotating flagellar filament to adjust swimming velocity, transcription, and physiology for optimal fitness in different host lower intestinal niches.
-
Journal articleDe Chiara A, Giachino C, Pirillo MF, et al., 2026,
Broadly cross-reactive mRNA COVID-19 vaccine encoding trimeric RBDs and NSP12 mitigates immune imprinting.
, Mol Ther Nucleic Acids, Vol: 37, ISSN: 2162-2531The continuous evolution of SARS-CoV-2 variants, driven by mutations in the spike protein undermines viral recognition by antibodies elicited through prior infection or vaccination with the ancestral Wuhan strain. Original antigenic sin of SARS-CoV-2 ancestral virus or vaccine led to a weakened neutralizing antibody response against successive variants upon administration of an updated vaccine. On the contrary, T cells retain cross-reactivity thanks to the high density of conserved epitopes. We designed mRNA vaccines encoding single-chain heterotrimers of the receptor-binding domain (RBD) natural variants of interest (VOI), (RBD-VOI) and of phylogenetically informed consensus representing the major variant lineages RBD-consensus (RBD-Cons). We demonstrate a broad neutralizing activity against omicron subvariants and mitigated immune imprinting when RBD-Cons was used as a booster after conventional Wuhan spike priming. To enhance cellular immunity, we designed a second mRNA vaccine component encoding the viral polymerase NSP12 able to induce a cross-reactive T cell response to be combined with the heterotrimeric RBD vaccine. Our results offer a rational strategy for next-generation, imprinting-resistant vaccines.
-
Journal articleOliver TJ, Elias E, Consoli G, et al., 2026,
Far-red chlorophyll d clusters extend photosystem I absorption toward the red limit.
, Sci Adv, Vol: 12Oxygenic photosynthesis is usually limited to visible light, but the marine cyanobacterium Acaryochloris marina pushes this boundary by harvesting far-red photons with chlorophyll d. The best-studied strain, MBIC11017, unexpectedly lacks low-energy chlorophylls ("red forms") in photosystem I, limiting absorption beyond 740 nanometers. Here, we show that another strain, A. marina NIES-2412, has evolved a strategy to absorb far-red photons up to 760 nanometers. Combining time-resolved fluorescence spectroscopy with cryo-electron microscopy at 2.64-angstrom resolution, we identify two distinct classes of chlorophyll d red forms in its photosystem I. One class originates from classical charge-transfer-exciton mixing, while the other arises purely from excitonic interactions. Mapping all 96 chlorophylls d reveals the precise pigments responsible for these far-red states. We also uncover a previously unreported subunit, PsaX2, which stabilizes the photosystem I complex and shapes pigment geometry and energetics to enable the formation of red forms. Last, we show that the protein modifications responsible for binding and tuning these red forms are widespread across the Acaryochloris genus but not within the model MBIC11017 strain. Far-red photons lie close to the energetic limit of oxygenic photosynthesis; their efficient use therefore requires fine-tuning of the photosynthetic machinery. To our knowledge, our findings provide the structural and mechanistic basis of one of the most red-shifted photosystem I complexes identified to date, highlighting a distinct adaptive strategy in far-red light environments and offering design principles for extending photosynthesis in crops into the infrared.
-
Journal articleBiswas P, Mishra V, Sanchez-Garrido J, et al., 2026,
Context-dependent epithelial and immune programs shape intestinal resilience or vulnerability following prior colitis
, Cellular and Molecular Gastroenterology and Hepatology (CMGH), ISSN: 2352-345XBackground & AimsPrior intestinal inflammation can leave durable immune and epithelial alterations, yet how these changes influence responses to subsequent injury remains unclear. Infectious and sterile colitis share core features, including barrier disruption and cytokine secretion. We therefore investigated whether the nature of the initial inflammatory event shapes protection or susceptibility during later intestinal insult.MethodsWe used reciprocal mouse models of Citrobacter rodentium (CR) infection and dextran sodium sulphate (DSS)-induced colitis to define how prior infectious versus sterile colitis shapes secondary disease. Barrier integrity, immune cell populations, cytokine production, and susceptibility to wild-type and CR mutants that cause limited epithelial barrier disruption were assessed.ResultsMice recovered from CR infection were protected against DSS-induced colitis, displaying reduced weight loss, preserved epithelial architecture, and lower inflammatory pathology. This protection required type III secretion system effector-mediated epithelial injury during primary infection and was associated with sustained IL-17A signalling, which contributed to the protective phenotype. In contrast, mice recovered from DSS colitis exhibited persistent epithelial barrier defects, chronic colonic neutrophilia, and heightened susceptibility to CR infection despite elevated IL-17A. Infection with CR mutants that cause minimal epithelial damage still resulted in severe disease in DSS-experienced mice, indicating that unresolved epithelial barrier dysfunction is a major contributor to vulnerability.ConclusionsThe nature of the primary colitis is associated with distinct epithelial and immune programs that persist beyond resolution of inflammation. Infectious colitis is associated with a protective mucosal state where IL-17A is a key contributor in a broader protective response, whereas sterile colitis is associated with persistent epithelial barrier dysfunction
-
Journal articleDewan D, Wang Y, De Simone A, et al., 2026,
Energy Landscape Analysis of Membrane Proteins Using NMR-Based Hybrid Restraint Potentials.
, J Chem Theory Comput, Vol: 22, Pages: 5827-5837Most biomolecular simulations depend on the quality of empirical force fields, and the use of hybrid restraint potentials has emerged as a promising approach. In this contribution, we extend the application of hybrid potentials to membrane proteins by developing optimized restraints derived from experimentally determined NMR data. NMR chemical shift, chemical shift anisotropy, dipolar coupling, and NOE distance information are combined with appropriately weighted empirical force fields to study two transmembrane systems, namely sarcolipin and phospholamban. To remedy the problems of rare events and broken ergodicity, the energy landscape framework, including basin-hopping global optimization and discrete path sampling, is employed for exploring the underlying energy landscapes. Much of the appeal of the hybrid potential approach is the ability to study membrane proteins in the absence of conventional explicit or implicit solvent and lipid molecules, thereby simplifying the sampling of complex biomolecular conformational spaces. Our results suggest that the hybridization of NMR constraints as penalty energies with empirical force fields improves global optimization and energy landscape analysis by excluding experimentally incompatible structures.
-
Journal articleBennison DJ, Chaudhary I, Chaudhuri D, et al., 2026,
GBP1 recruitment to actin-rich pedestals of extracellular Gram-negative bacteria promotes pyroptosis
, EMBO Journal, ISSN: 0261-4189The IFNγ-induced GTPase guanylate-binding protein 1 (GBP1) binds to lipopolysaccharide (LPS) on cytosolic gram-negative bacteria and promotes pyroptosis via the recruitment and activation of caspase-4 on the bacterial outer membrane. Enteropathogenic and enterohaemorrhagic Escherichia coli (EPEC and EHEC, respectively) are extracellular pathogens that adhere to host cells and stimulate dense actin polymerisation underneath their attachment sites, generating structures described as actin-rich pedestals. Here, we show that GBP1 traffics to actin-rich pedestals in human cells infected with EPEC or EHEC in vitro and mouse colonocytes infected with the EPEC-like murine pathogen Citrobacter rodentium in vivo. GBP1 promotes caspase-4 recruitment to actin-rich pedestals, leading to pyroptosis and IL-18 release. GBP1 mutants defective in LPS coatomer formation also localise to EPEC pedestals. A novel assay that mimics pathogenic effector activity reveals GBP1 recruitment to sterile actin polymerisation sites. We conclude that cytosolic GBP1 is mobilised to sites of pathogen-induced actin remodelling independently of LPS. Our study establishes that GBP1 not only operates as a pattern-recognition receptor but also orchestrates effector-triggered immunity against pathogens that hijack the actin cytoskeleton.
This data is extracted from the Web of Science and reproduced under a licence from Thomson Reuters. You may not copy or re-distribute this data in whole or in part without the written consent of the Science business of Thomson Reuters.