Publications

@article{noKey,

title = {Heterologous production, purification and crystallization of 24C-sterol methyltransferase from Candida albicans},

author = {S. Mpontshane, Nokwanda},

url = {https://www.biorxiv.org/content/10.1101/2025.02.05.636639v1.abstract},

doi = {https://doi.org/10.1101/2025.02.05.636639},

year  = {2025},

date = {2025-02-05},

abstract = {Candida albicans is a critical priority fungal pathogen causing invasive fungal infections with high mortality rates in immunocompromised patients. The increasing fungal infection rate and resistance of fungal pathogens to existing antifungal treatments have emphasized the need for the development of novel antifungal medicine. The ergosterol biosynthesis pathway has been a successful target for antifungal compounds, but many enzymatic steps remain unexplored. 24C-sterol methyltransferase (24C-SMT) catalyzes a critical fungal-specific step in ergosterol biosynthesis. When 24C-SMT is disrupted, fungal pathogens are sensitized to temperature, various inhibitors, and antifungals, and a loss of virulence can be observed. In this study, five 24C-SMT variants with different lengths of N-termini were heterologously produced in Escherichia coli and three were purified to near-homogeneity with immobilized metal-affinity and size-exclusion chromatography. N-terminally truncated C. albicans 24C-SMT was utilized for crystallization trials due to its increased stability and higher purity compared to the full-length protein. 24C-SMT crystals were obtained in the presence of Sadenosyl-homocysteine, but diffracted to low resolution. Therefore, we established a starting point for 24C-SMT crystallization by providing an optimized protocol for heterologous 24C-SMT production, purification and initial crystallization conditions, which could be used for further downstream crystallographic studies.},

keywords = {FORMULATOR},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Beyond Barriers, Big Crystallisation Hurdles: Atropisomerism in bRo5 Compounds Explored by Computational and NMR Studies},

author = {Angelos Stamos, Nikolaos},

url = {https://chemrxiv.org/engage/chemrxiv/article-details/679a3f956dde43c908d979c7},

doi = {https://doi.org/10.26434/chemrxiv-2025-rz4q6},

year  = {2025},

date = {2025-02-03},

abstract = {Crystallisation and stereochemical stability are pivotal factors in pharmaceutical development, particularly for complex beyond Rule of 5 (bRo5) compounds. In this study, we explore the intricate interplay between atropisomerism and crystallisation using two model bRo5 compounds, namely ACBI1 and BI201335, both violating three of four Lipinski’s rules. One of the tool compounds exhibits Class 2 atropisomeric behaviour and the other devoid of it. A diverse array of crystallisation methods—including solution-phase crystallisation, cocrystallisation, and salt formation—was applied, revealing the critical role of atropisomerism induced stereochemistry in polymorphism and nucleation outcomes. In-silico torsion profile calculations and NMR studies were employed to elucidate the rotational energy barriers and confirm the presence or absence of atropisomerism.  This comprehensive analysis highlights the significance of understanding stereochemical phenomena like atropisomerism in designing and developing bRo5 compounds. By integrating advanced analytical techniques and crystallisation strategies, this work provides novel insights into tailoring pharmaceutical properties for nextgeneration therapeutics.},

keywords = {ROCKIMAGER},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {COVID-19 Breakthrough Infections in Immune-Mediated Inflammatory Diseases: Data from the SUCCEED (Safety and Immunogenicity of COVID-19 Vaccines in Systemic Autoimmune-Mediated Inflammatory Diseases) Study},

author = {Tan, Jeremiah, Bernatsky, Sasha, et al.},

url = {https://www.mdpi.com/2076-393X/13/2/104},

doi = {https://doi.org/10.3390/vaccines13020104},

year  = {2025},

date = {2025-01-22},

abstract = {ackground: The Safety and Immunogenicity of COVID-19 Vaccines in Systemic Autoimmune-Mediated Inflammatory Diseases (SUCCEED) study was created to better understand COVID-19 vaccination in immune-mediated inflammatory disease (IMID). Knowing the frequency of COVID-19 breakthrough infections is important, particularly in IMID. Our objective was to assess these events in IMID. Methods: We prospectively studied IMID participants who had received ≥three COVID-19 vaccine doses. Individuals provided saliva samples monthly (September 2022 to August 2023). These were evaluated by polymerase chain reaction (PCR) for SARS-CoV-2. We also assessed antibodies against SARS-CoV-2 (anti-spike, SmT1, receptor binding domain, RBD, and nucleocapsid, NP) based on dried blood spots. Multivariable general estimating equation regression produced odd ratios (OR) for PCR SARS-CoV-2 positivity, related to demographics, immunosuppressives, and antibody levels. Results: Diagnoses included rheumatoid arthritis RA (N = 161, 44% of the total), systemic lupus, psoriatic arthritis, spondylarthritis, vasculitis, systemic sclerosis, and inflammatory bowel disease. Of the 366 participants, most were taking immunosuppressive medication. Of 1266 saliva samples, 56 (5.1%) were positive for SARS-CoV-2 on PCR. Higher anti-SmT1 antibodies were inversely associated with SARS-CoV-2 detection on PCR (adjusted OR 0.66, 95% confidence interval 0.45–0.97). Antibodies to SmT1, RBD, and NP were correlated and thus could not be included in a single model, but when anti-RBD was used in place of anti-SmT1, the results were similar. No other factor (including prior COVID-19 infection) was clearly associated with SARS-CoV-2 detection. Conclusions: This is the first study of SARS-CoV-2 in a large prospective cohort of triple (or more) vaccinated individuals with IMIDs. Anti-SmT1 antibodies appeared to be protective against later SARS-CoV-2 positivity, although recent past infection was not clearly related. This suggests the importance of maintaining robust vaccine-induced immunity through vaccination in IMID.},

keywords = {MANTIS},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Rapid Development of High Concentration Protein Formulation Driven by High-Throughput Technologies},

author = {Xin et al., Lun},

url = {https://link.springer.com/article/10.1007/s11095-024-03801-3},

doi = {https://doi.org/10.1007/s11095-024-03801-3},

year  = {2025},

date = {2025-01-17},

abstract = {Background High concentration protein formulation (HCPF) development needs to balance protein stability attributes such

as conformational/colloidal stability, chemical stability, and solution properties such as viscosity and osmolality.

Methodology A three-phase design is established in this work. In Phase 1, conformational and colloidal stability are measured by 384-well-based high-throughput (HT) biophysical screening while viscosity reduction screening is performed with HT viscosity screening. Collectively, the biophysical and viscosity screening data are leveraged to design the phase 2 of short-term stability study, executed using 96-well plates under thermal and freeze/thaw stresses. In phase 2, samples are analyzed by stability-indicating assays and processed with pair-wise Student’s t-test analyses to choose the final formulations. In phase 3, the final formulations are then confirmed through a one-month accelerated stability in glass vials.

Results Using a model antibody A (mAb-A), the initial HT screening successfully established the 384-well based platform.

A lead formulation was chosen from the second round based on statistical analyses and subsequently tested against the commercial

formulation of mAb-A as a control. Compared to the control, the lead formulation reduced the viscosity of mAb-A by 30% and decreased subvisible particles after thermal stress by 80%.

Conclusions HT biophysical screening in 384-well plates was demonstrated to effectively guide the rational design of a high-throughput stability screening study using 96-well plates. This platform enables the identification of a high concentration formulation within seven weeks within the first two phases of study that strategically balance stability with solution

properties, thus achieving a rapid development of HCPF.},

keywords = {µPULSE},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {From dimer to tetramer: the evolutionary trajectory of C4 photosynthetic-NADP-ME oligomeric state in Poaceae},

author = {M. Böhm, Jonas},

url = {https://www.biorxiv.org/content/10.1101/2025.01.05.631420v1.abstract},

doi = {https://doi.org/10.1101/2025.01.05.631420},

year  = {2025},

date = {2025-01-06},

abstract = {The C4 carbon concentrating mechanism relies on specialized enzymes that have evolved unique expression patterns and biochemical properties distinct to their ancestral housekeeping forms. In maize and sorghum, the evolution of C4-NADP-malic enzyme (C4-NADP-ME) involved gene duplication and neofunctionalization, leading to the emergence of two plastidic isoforms: C4-NADP-ME and nonC4-NADP-ME, each with distinct kinetic and structural features. While C4-NADP-ME functions primarily as a tetramer, nonC4-NADP-ME exists in an equilibrium between dimeric and tetrameric forms, favoring the dimer in solution. This study shows which evolutionary changes in amino acid sequences influence the structure and function of these isoforms. By integrating X-ray crystallography, cryo-electron microscopy, computational molecular modeling and targeted biochemical analysis of mutant and truncated protein variants, we identify crucial roles for the N- and C-terminal regions and specific amino acid residues in governing isoform oligomerization. Our results reveal that the N-terminal region is essential for stabilizing the dimeric form of nonC4-NADP-ME, whereas specific adaptive substitutions and interactions with the C-terminal region enhance the stability of the tetrameric state characteristic of the C4-adapted isoform. We propose that differences in the N-terminal domain between the C4 and nonC4 isoforms reflect distinct selective pressures, which have driven their evolutionary divergence to fulfill specialized cellular functions.},

keywords = {ROCKIMAGER},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Discovery and mechanism of a highly selective, antifungal acetyl CoA synthetase inhibitor},

author = {Krysan, Damian},

url = {https://www.researchsquare.com/article/rs-5619443/v1},

doi = {https://doi.org/10.21203/rs.3.rs-5619443/v1},

year  = {2025},

date = {2025-01-01},

abstract = {Acetyl CoA synthetases (ACS) have emerged as drug targets for the treatment of cancer, metabolic diseases as well as fungal and parasitic infections. Although a variety of small molecule ACS inhibitors have been discovered, the systematic optimization of these molecules has been slowed by a lack of structural information regarding their mechanism of inhibition. Through a chemical genetic-based, synthetic lethal screen of the human fungal pathogen Cryptococcus neoformans, we identified an isoxazole-based ACS inhibitor with antifungal activity and exquisite selectivity for the C. neoformans Acs1 relative to human ACSS2 as well as other fungal ACSs. Xray crystallographic characterization of the isoxazole-CnAcs1 complex revealed that the isoxazole functions as an acetyl CoA mimic and occupies both the acetyl- and CoA-binding sites of CnAcs1. Consistent with this novel mode of inhibition, the isoxazoles display uncompetitive inhibition kinetics that are similar to antimalarial ACS inhibitors also proposed to target the CoA binding site. Consequently, these data provide structural and mechanistic insights into the remarkable selectivity of Acetyl CoA pocket-targeting ACS inhibitors. In addition, these data provide strong proof-of-principle that targeting fungal and parasitic ACSs for the development of novel anti-infectives can be achieved with high selectivity and, thereby, low host toxicity.},

keywords = {NT8},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {RNA-dependent RNA polymerase of predominant human norovirus forms liquid-liquid phase condensates as viral replication factories},

author = {Kaundal, Soni},

url = {https://www.science.org/doi/full/10.1126/sciadv.adp9333},

doi = {https://doi.org/10.1126/sciadv.adp9333},

year  = {2024},

date = {2024-12-20},

abstract = {Many viral proteins form biomolecular condensates via liquid-liquid phase separation (LLPS) to support viral replication and evade host antiviral responses, and thus, they are potential targets for designing antivirals. In the case of nonenveloped positive-sense RNA viruses, forming such condensates for viral replication is unclear and less understood. Human noroviruses (HuNoVs) are positive-sense RNA viruses that cause epidemic and sporadic gastroenteritis worldwide. Here, we show that the RNA-dependent RNA polymerase (RdRp) of pandemic GII.4 HuNoV forms distinct condensates that exhibit all the signature properties of LLPS with sustained polymerase activity and the capability of recruiting components essential for viral replication. We show that such condensates are formed in HuNoV-infected human intestinal enteroid cultures and are the sites for genome replication. Our studies demonstrate the formation of phase-separated condensates as replication factories in a positive-sense RNA virus, which plausibly is an effective mechanism to dynamically isolate RdRp replicating the genomic RNA from interfering with the ribosomal translation of the same RNA.},

keywords = {ROCKIMAGER},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Enhanced identification of small molecules binding to hnRNPA1 via cryptic pockets mapping coupled with X-Ray fragment screening},

author = {Dunnett, Louise},

url = {https://www.biorxiv.org/content/10.1101/2024.12.17.628909v1.abstract},

doi = {https://doi.org/10.1101/2024.12.17.628909},

year  = {2024},

date = {2024-12-18},

abstract = {The human heterogeneous nuclear ribonucleoprotein (hnRNP) A1 is a prototypical RNA-binding protein essential in regulating a wide range of post-transcriptional events in cells. As a multifunctional protein with a key role in RNA metabolism, deregulation of its functions has been linked to neurodegenerative diseases, tumour aggressiveness and chemoresistance, which has fuelled efforts to develop novel therapeutics that modulates its RNA binding activities. Here, using a combination of Molecular Dynamics (MD) simulations and graph neural network pockets predictions, we showed that hnRNPA1 N-terminal RNA binding domain (UP1) contains several cryptic pockets capable of binding small molecules. To identify chemical entities for development of potent drug candidates and experimentally validate identified druggable hotspots, we carried out a large fragment screening on UP1 protein crystals. Our screen identified 36 hits which extensively samples UP1 functional regions involved in RNA recognition and binding, as well as mapping hotspots onto novel protein interaction surfaces. We observed a wide range of ligand-induced conformational variation, by stabilisation of dynamic protein regions. Our high-resolution structures, the first of an hnRNP in complex with a fragment or small molecule, provides rapid routes for the rational development of a range of different inhibitors and chemical tools for studying molecular mechanisms of hnRNPA1 mediated splicing regulation.},

keywords = {ROCKIMAGER},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Engineering and structures of Crimean-Congo hemorrhagic fever virus glycoprotein complexes},

author = {McFadden, Elizabeth},

url = {https://www.cell.com/cell/fulltext/S0092-8674(24)01325-4},

doi = {https://doi.org/10.1016/j.cell.2024.11.008},

year  = {2024},

date = {2024-12-18},

abstract = {Crimean-Congo hemorrhagic fever virus (CCHFV) is a tickborne virus that can cause severe disease in humans with case fatality rates of 10%–40%. Although structures of CCHFV glycoproteins GP38 and Gc have provided insights into viral entry and defined epitopes of neutralizing and protective antibodies, the structure of glycoprotein Gn and its interactions with GP38 and Gc have remained elusive. Here, we use structure-guided protein engineering to produce a stabilized GP38-Gn-Gc heterotrimeric glycoprotein complex (GP38-GnH-DS-Gc). A cryo-electron microscopy (cryo-EM) structure of this complex provides the molecular basis for GP38’s association on the viral surface, reveals the structure of Gn, and demonstrates that GP38-Gn restrains the Gc fusion loops in the prefusion conformation, facilitated by an N-linked glycan attached to Gn. Immunization with GP38-GnH-DS-Gc conferred 40% protection against lethal IbAr10200 challenge in mice. These data define the architecture of a GP38-Gn-Gc protomer and provide a template for structure-guided vaccine antigen development.},

keywords = {NT8},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Biophysical Characterization of a Novel Phosphopentomutase from the Hyperthermophilic Archaeon Thermococcus kodakarensis},

author = {Naz, Zahra},

url = {https://www.mdpi.com/1422-0067/25/23/12893},

doi = {https://doi.org/10.3390/ijms252312893},

year  = {2024},

date = {2024-11-30},

abstract = {Phosphopentomutases catalyze the isomerization of ribose 1-phosphate and ribose 5-phosphate. Thermococcus kodakarensis, a hyperthermophilic archaeon, harbors a novel enzyme (PPMTk) that exhibits high homology with phosphohexomutases but has no significant phosphohexomutase activity. Instead, PPMTk catalyzes the interconversion of ribose 1-phosphate and ribose 5-phosphate. Here, we report biophysical analysis, crystallization, and three-dimensional structure determination of PPMTk by X-ray diffraction at 2.39 Å resolution. The solved structure revealed a novel catalytic motif, unique to PPMTk, which makes this enzyme distinct from the homologous counterparts. We postulate that this novel catalytic motif may enable PPMTk to isomerize phosphopentose instead of phosphohexose. To the best of our knowledge, this is the first biophysical and structural analysis of a phosphopentomutase from hyperthermophilic archaea.},

keywords = {ROCKIMAGER},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Allosteric substrate release by a sialic acid TRAP transporter substrate binding protein},

author = {Schneberger, Niels},

url = {https://www.nature.com/articles/s42003-024-07263-6},

doi = {https://doi.org/10.1038/s42003-024-07263-6},

year  = {2024},

date = {2024-11-23},

abstract = {The tripartite ATP-independent periplasmic (TRAP) transporters enable Vibrio cholerae and Haemophilus influenzae to acquire sialic acid, aiding their colonization of human hosts. This process depends on SiaP, a substrate-binding protein (SBP) that captures and delivers sialic acid to the transporter. We identified 11 nanobodies that bind specifically to the SiaP proteins from H. influenzae (HiSiaP) and V. cholerae (VcSiaP). Two nanobodies inhibited sialic acid binding. Detailed structural and biophysical studies of one nanobody-SBP complex revealed an allosteric inhibition mechanism, preventing ligand binding and releasing pre-bound sialic acid. A hydrophobic surface pocket of the SBP is crucial for the allosteric mechanism and for the conformational rearrangement that occurs upon binding of sialic acid to the SBP. Our findings provide new clues regarding the mechanism of TRAP transporters, as well as potential starting points for novel drug design approaches to starve these human pathogens of important host-derived molecules.},

keywords = {ROCKIMAGER},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Viral sequence determines HLA-E-restricted T cell recognition of hepatitis B surface antigen},

author = {Murugesan, Gavuthami},

url = {https://www.nature.com/articles/s41467-024-54378-9},

doi = {https://doi.org/10.1038/s41467-024-54378-9},

year  = {2024},

date = {2024-11-22},

abstract = {The non-polymorphic HLA-E molecule offers opportunities for new universal immunotherapeutic approaches to chronic infectious diseases. Chronic Hepatitis B virus (HBV) infection is driven in part by T cell dysfunction due to elevated levels of the HBV envelope (Env) protein hepatitis B surface antigen (HBsAg). Here we report the characterization of three genotypic variants of an HLA-E-binding HBsAg peptide, Env371-379, identified through bioinformatic predictions and verified by biochemical and cellular assays. Using a soluble affinity-enhanced T cell receptor (TCR) (a09b08)-anti-CD3 bispecific molecule to probe HLA-E presentation of the Env371-379 peptides, we demonstrate that only the most stable Env371-379 variant, L6I, elicits functional responses to a09b08-anti-CD3-redirected polyclonal T cells co-cultured with targets expressing endogenous HBsAg. Furthermore, HLA-E-Env371-379 L6I-specific CD8+ T cells are detectable in HBV-naïve donors and people with chronic HBV after in vitro priming. In conclusion, we provide evidence for HLA-E-mediated HBV Env peptide presentation, and highlight the effect of viral mutations on the stability and targetability of pHLA-E molecules.},

keywords = {ROCKIMAGER},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Single-cell sequencing of full-length transcripts and T-cell receptors with automated high-throughput Smart-seq3},

author = {Chuang, Hsiu-Chun, Ruidong, Li, Li, Li, Kai-Hui, Sun},

url = {https://bmcgenomics.biomedcentral.com/articles/10.1186/s12864-024-11036-0},

doi = {https://doi.org/10.1186/s12864-024-11036-0},

year  = {2024},

date = {2024-11-21},

abstract = {We developed an automated high-throughput Smart-seq3 (HT Smart-seq3) workflow that integrates best practices and an optimized protocol to enhance efficiency, scalability, and method reproducibility. This workflow consistently produces high-quality data with high cell capture efficiency and gene detection sensitivity. In a rigorous comparison with the 10X platform using human primary CD4 + T-cells, HT Smart-seq3 demonstrated higher cell capture efficiency, greater gene detection sensitivity, and lower dropout rates. Additionally, when sufficiently scaled, HT Smart-seq3 achieved a comparable resolution of cellular heterogeneity to 10X. Notably, through T-cell receptor (TCR) reconstruction, HT Smart-seq3 identified a greater number of productive alpha and beta chain pairs without the need for additional primer design to amplify full-length V(D)J segments, enabling more comprehensive TCR profiling across a broader range of species. Taken together, HT Smart-seq3 overcomes key technical challenges, offering distinct advantages that position it as a promising solution for the characterization of single-cell transcriptomes and immune repertoires, particularly well-suited for low-input, low-RNA content samples.},

keywords = {MANTIS},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Engineering of soluble bacteriorhodopsin},

author = {Nikolaev, Andrey},

url = {https://www.biorxiv.org/content/10.1101/2024.11.20.624543v1.abstract},

doi = {https://doi.org/10.1101/2024.11.20.624543},

year  = {2024},

date = {2024-11-21},

abstract = {Bacteriorhodopsin is a seven-helical light-driven proton pump and a model membrane protein. Here, we report engineering of soluble analogues of bacteriorhodopsin, NeuroBRs, which bind retinal and photocycle under illumination. We also report the crystallographic structure of NeuroBR_A, determined at anisotropic resolution reaching 1.76 Å, that reveals a conserved chromophore binding pocket and tertiary structure. Our results highlight the power of modern protein engineering approaches and pave the way towards wider development of molecular tools derived from membrane proteins.},

keywords = {NT8},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Broadening alloselectivity of T cell receptors by structure guided engineering},

author = {Karuppiah, Vijaykumar},

url = {https://www.nature.com/articles/s41598-024-75140-7#Abs1},

doi = {https://doi.org/10.1038/s41598-024-75140-7},

year  = {2024},

date = {2024-11-06},

abstract = {Specificity of a T cell receptor (TCR) is determined by the combination of its interactions to the peptide and human leukocyte antigen (HLA). TCR-based therapeutic molecules have to date targeted a single peptide in the context of a single HLA allele. Some peptides are presented on multiple HLA alleles, and by engineering TCRs for specific recognition of more than one allele, there is potential to expand the targetable patient population. Here, as a proof of concept, we studied two TCRs, S2 and S8, binding to the PRAME peptide antigen (ELFSYLIEK) presented by HLA alleles HLA-A*03:01 and HLA-A*11:01. By structure-guided affinity maturation targeting a specific residue on the HLA surface, we show that the affinity of the TCR can be modulated for different alleles. Using a combination of affinity maturation and functional T cell assay, we demonstrate that an engineered TCR can target the same peptide on two different HLA alleles with similar affinity and potency. This work highlights the importance of engineering alloselectivity for designing TCR based therapeutics suitable for differing global populations.},

keywords = {ROCKIMAGER},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {A bacterial immunity protein directly senses two disparate phage proteins},

author = {Zhang, Tong},

url = {https://www.nature.com/articles/s41586-024-08039-y},

doi = {https://doi.org/10.1038/s41586-024-08039-y},

year  = {2024},

date = {2024-10-16},

abstract = {Eukaryotic innate immune systems use pattern recognition receptors to sense infection by detecting pathogen-associated molecular patterns, which then triggers an immune response. Bacteria have similarly evolved immunity proteins that sense certain components of their viral predators, known as bacteriophages1,2,3,4,5,6. Although different immunity proteins can recognize different phage-encoded triggers, individual bacterial immunity proteins have been found to sense only a single trigger during infection, suggesting a one-to-one relationship between bacterial pattern recognition receptors and their ligands7,8,9,10,11. Here we demonstrate that the antiphage defence protein CapRelSJ46 in Escherichia coli can directly bind and sense two completely unrelated and structurally different proteins using the same sensory domain, with overlapping but distinct interfaces. Our results highlight the notable versatility of an immune sensory domain, which may be a common property of antiphage defence systems that enables them to keep pace with their rapidly evolving viral predators. We found that Bas11 phages harbour both trigger proteins that are sensed by CapRelSJ46 during infection, and we demonstrate that such phages can fully evade CapRelSJ46 defence only when both triggers are mutated. Our work shows how a bacterial immune system that senses more than one trigger can help prevent phages from easily escaping detection, and it may allow the detection of a broader range of phages. More generally, our findings illustrate unexpected multifactorial sensing by bacterial defence systems and complex coevolutionary relationships between them and their phage-encoded triggers.},

keywords = {NT8},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {A single Leishmania adenylate forming enzyme of the ANL superfamily generates both acetyl- and acetoacetyl-CoA},

author = {J. Jezewski, Andrew},

url = {https://www.jbc.org/article/S0021-9258(24)02381-0/fulltext},

doi = {https://doi.org/10.1016/j.jbc.2024.107879},

year  = {2024},

date = {2024-10-09},

abstract = {Leishmania, a protozoan parasite, is responsible for significant morbidity and mortality worldwide, manifesting as cutaneous, mucocutaneous, and visceral leishmaniasis. These diseases pose a substantial burden, especially in impoverished regions with limited access to effective medical treatments. Current therapies are toxic, have low efficacy, and face growing resistance. Understanding the metabolic pathways of Leishmania, particularly those differing from its host, can unveil potential therapeutic targets. In this study, we investigated the acetyl-CoA synthetase (ACS) enzyme from Leishmania infantum (LiAcs1), which, unlike many organisms, also exhibits acetoacetyl-CoA synthetase (KBC) activity. This dual functionality is unique among ANL superfamily enzymes and crucial for the parasite's reliance on leucine catabolism, energy production and sterol biosynthesis. Our biochemical characterization of LiAcs1 revealed its ability to utilize both acetate and acetoacetate substrates. Additionally, LiAcs1 displayed a distinct CoA substrate inhibition pattern, partially alleviated by acetoacetate. Structural analysis provided insights into the substrate binding flexibility of LiAcs1, highlighting a more promiscuous substrate pocket compared to other ACS or KBC-specific enzymes. Substrate mimetics elucidated its ability to accommodate both small and large AMP-ester derivatives, contributing to its dual ACS/KBC functionality. These findings not only advance our understanding of Leishmania metabolism but also present LiAcs1 as a promising drug target. The dual functionality of LiAcs1 underscores the potential for developing selective inhibitors that could disrupt critical metabolic pathways across Leishmania spp. as it appears this enzyme is highly conserved across this genus. This paves the way for developing novel effective treatments against this devastating disease.},

keywords = {NT8},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Targeting RSV-neutralizing B cell receptors with anti-idiotypic antibodies},

author = {C. Scharffenberger, Samuel},

url = {https://www.cell.com/cell-reports/fulltext/S2211-1247(24)01162-8},

doi = {https://doi.org/10.1016/j.celrep.2024.114811},

year  = {2024},

date = {2024-10-08},

abstract = {Respiratory syncytial virus (RSV) causes lower respiratory tract infections with significant morbidity and mortality at the extremes of age. Vaccines based on the viral fusion protein are approved for adults over 60, but infant protection relies on passive immunity via antibody transfer or maternal vaccination. An infant vaccine that rapidly elicits protective antibodies would fulfill a critical unmet need. Antibodies arising from the VH3-21/VL1-40 gene pairing can neutralize RSV without the need for affinity maturation, making them attractive to target through vaccination. Here, we develop an anti-idiotypic monoclonal antibody (ai-mAb) immunogen that is specific for unmutated VH3-21/VL1-40 B cell receptors (BCRs). The ai-mAb efficiently engages B cells with bona fide target BCRs and does not activate off-target non-neutralizing B cells, unlike recombinant pre-fusion (preF) protein used in current RSV vaccines. These results establish proof of concept for using an ai-mAb-derived vaccine to target B cells hardwired to produce RSV-neutralizing antibodies.},

keywords = {NT8},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Structure-Based Engineering of Monoclonal Antibodies for Improved Binding to Counteract the Effects of Fentanyl and Carfentanil},

author = {Rodarte, Justas},

url = {https://pubs.acs.org/doi/full/10.1021/acsomega.4c06617},

doi = {https://doi.org/10.1021/acsomega.4c06617},

year  = {2024},

date = {2024-10-07},

abstract = {The opioid overdose epidemic is a growing and evolving public health crisis fueled by the widespread presence of fentanyl and fentanyl analogues (F/FAs) in both street mixtures and counterfeit pills. To expand current treatment options, drug-targeting monoclonal antibodies (mAbs) offer a viable therapeutic for both pre- and postexposure clinical scenarios. This study reports the isolation, in vitro characterization, and in vivo efficacy of two murine mAb families targeting fentanyl, carfentanil, or both. Because humanization of the mAbs by CDR grafting negatively impacted affinity for both fentanyl and carfentanil, crystal structures of mAbs in complex with fentanyl or carfentanil were analyzed to identify key residues involved in ligand binding in murine versus humanized structures, and site-directed mutagenesis was used to verify their functional importance. The structural analysis identified a framework residue, Tyr36, present in the murine germline sequence of two mAbs, which was critical for binding to fentanyl and carfentanil. These studies emphasize the importance of structural considerations in mAb engineering to optimize mAbs targeting small molecules including opioids and other drugs of public health interest.},

keywords = {NT8},

pubstate = {published},

tppubtype = {article}

}

@article{noKey,

title = {Structural studies of β-glucosidase from the thermophilic bacterium Caldicellulosiruptor saccharolyticus},

author = {I. Sotiropoulou, Anastasia},

url = {https://journals.iucr.org/d/issues/2024/10/00/gm5108/index.html},

doi = {https://doi.org/10.1107/S2059798324009252},

year  = {2024},

date = {2024-10-01},

abstract = {β-Glucosidase from the thermophilic bacterium Caldicellulosiruptor saccharo­lyticus (Bgl1) has been denoted as having an attractive catalytic profile for various industrial applications. Bgl1 catalyses the final step of in the decomposition of cellulose, an unbranched glucose polymer that has attracted the attention of researchers in recent years as it is the most abundant renewable source of reduced carbon in the biosphere. With the aim of enhancing the thermostability of Bgl1 for a broad spectrum of biotechnological processes, it has been subjected to structural studies. Crystal structures of Bgl1 and its complex with glucose were determined at 1.47 and 1.95 Å resolution, respectively. Bgl1 is a member of glycosyl hydrolase family 1 (GH1 superfamily, EC 3.2.1.21) and the results showed that the 3D structure of Bgl1 follows the overall architecture of the GH1 family, with a classical (β/α)8 TIM-barrel fold. Comparisons of Bgl1 with sequence or structural homologues of β-glucosidase reveal quite similar structures but also unique structural features in Bgl1 with plausible functional roles.},

keywords = {ROCKIMAGER},

pubstate = {published},

tppubtype = {article}

}