Biology subjects

Pagel, K.

Publications and source records attributed to Pagel, K..

8 recordsLinked to original sources

bioRxiv2026

Glycosaminoglycans Promote Amyloid-β Aggregation via Multivalent, pH-Dependent Interactions

Glycosaminoglycans (GAGs) are polyanionic polysaccharides that co-localize with amyloid-{beta} (A{beta}) deposits in Alzheimers disease, yet their mechanistic contribution to A{beta} aggregation remains unclear. Here, we show that GAGs function as pH-responsive electrostatic scaffolds that selectively accelerate A{beta}(1-42) aggregation under mildly acidic, endosomal conditions but not at neutral extracellular pH. Combining experimental and computational approaches, we identify protonated N-terminal histidines as key determinants of GAG binding. Weak interactions between GAGs and the charged Nterminal region of A{beta} promote conformational rearrangements that bring peptides into proximity and expose adjacent hydrophobic aggregation-prone segments, thereby facilitating peptide clustering. Kinetic analyses reveal that aggregation is enhanced in a way consistent with an apparent increase in effective peptide concentration, accelerating nucleation without altering the dominant aggregation pathway. Systematic variation of GAG chain length and sulfation level further demonstrates that aggregation enhancement requires a threshold degree of multivalency, consistent with a clustering-driven mechanism. Together, these findings establish a framework in which pH-dependent electrostatic interactions with GAGs act as molecular triggers of amyloid nucleation, providing insight into how cellular microenvironments regulate the earliest stages of Alzheimers disease pathology.

biophysics↗

bioRxiv2025

HMGB1 B-box domain self-complexes promote protein-polyelectrolyte interactions.

HMGB1, a nuclear DNA-binding protein, can be secreted by activated immune cells or passively released from damaged cells. In such cases, HMGB1 functions as an alarmin that activates the immune system. Excessive inflammation may lead to pathogenesis, whereas this response can be dampened by polyanion binding, which impedes further receptor recognition. Moreover, HMGB1 is known to form liquid droplets in the cellular environment--a phase separation directly linked to its proper function. While the A-Box domain is believed to be primarily responsible for heparin binding due to its conserved binding site, the association and phase separation behavior of HMGB1 may be mediated by the B-box domain, owing to its extended hydrophobic regions. In this study, we first demonstrated that the B-box protein forms 30-nm large self-associates while maintaining its structure. Next, using molecularly sensitive EPR spectroscopy, we showed that the presence of these protein associations significantly enhances interactions with heparin. Notably, the local conformational changes induced by heparin are similar in both individual protein chains and their self-associated forms. To explain this effect, AlphaFold modeling was employed, revealing that the formation of protein multimers induces charge redistribution, resulting in an extended positively charged region that enhances electrostatic attraction to negatively charged polyanions, such as heparin.

biochemistry↗

bioRxiv2024

Structural Determinants of Mucins in Influenza Virus Inhibition: The Synergistic Role of Sialylated Glycans and Molecular Size

Mucins are heavily glycosylated proteins that play a crucial role in protecting mucosal surfaces against pathogens, including influenza viruses. This study investigates the antiviral properties of bovine submaxillary mucins (BSM) as a model for oral mucins against the influenza virus (A/H3N2 subtype), focusing on glycan composition and mucin size. BSM was purified, and characterized by proteomic and glycomic analysis and its antiviral efficacy was assessed after selective removal of sialic acids, N-glycans, or all glycans via enzymatic and chemical treatments. We employed virus binding and inhibition assays, including microscale thermophoresis (MST) and hemagglutination inhibition (HAI), to characterize processed mucins for structure activity correlations. Removal of sialic acids reduced BSMs antiviral activity by over 10-fold, while complete glycan removal abolished it entirely, highlighting sialylated O-glycans as critical for viral inhibition. N-glycan removal had minimal impact on antiviral efficacy. A size-dependent antiviral effect was observed: smaller mucin fragments ([~]50 and 330 kDa), which retained comparable O-glycosylation patterns, showed significantly reduced inhibition and viral binding affinity compared to intact BSM. These findings underscore the importance of mucin size and sialylated O-glycans in antiviral defense mechanisms against influenza. SignificanceThis study sheds light on the intrinsic antiviral properties of mucin from the bovine submaxillary gland, revealing the roles of glycans and mucins size in binding and inhibiting influenza virus. Our findings suggest a clear correlation between sialylated O-glycans and mucin size with the antiviral efficacy. Ultimately, we show that mucin-derived fragments retaining virus-binding capacity, with defined size and O-glycosylation patterns, can be isolated from mucin and could serve as versatile building blocks for designing next-generation antiviral biomaterials.

biochemistry↗

bioRxiv2024

Molecular and functional profiling unravels targetable vulnerabilities in colorectal cancer

While colorectal cancer (CRC) patients with microsatellite instability (MSI) respond well to immunotherapy those with microsatellite stable (MSS) tumors rely on conventional chemotherapy, often with poor outcomes. Both types frequently carry mutations in KRAS or BRAF proto-oncogenes, rendering them more resistant to treatment. New therapeutic biomarkers and treatments remain a clinical need, especially for MSS tumors. We performed whole exome and RNA-Sequencing from 28 tumors of the Athens Comprehensive Cancer Center CRC cohort, and molecularly characterized colorectal cancer patients based on their MSI status, SNVs/CNAs, and pathway/transcription factor activities at the individual patient level. Variants were classified using a new computational score for integrative cancer variant annotation and prioritization. Complementing this molecular data with public multi-omics datasets, we identified activation of transforming growth factor beta (TGF{beta}) signaling to be stronger activated in the MSS patients whereas JAK-STAT and MAPK molecular cascades were activated specifically in MSI. We unraveled mechanisms consistently perturbed in the transcriptional and mutational circuits and identified RUNX transcription factors as putative novel targets. Assessing the immunogenicity of CRC tumors in the context of RAS/RAF mutations and MSI/MSS status revealed a critical impact KRAS mutations have on immunogenicity particularly in the MSS patient subgroup, with implications for diagnosis and treatment.

cancer biology↗

bioRxiv2024

Enzymatic modulation of the pulmonary glycocalyx alters susceptibility to Streptococcus pneumoniae

The pulmonary epithelial glycocalyx is rich in glycosaminoglycans such as hyaluronan and heparan sulfate. Despite their presence, the precise role of these glycosaminoglycans in bacterial lung infections remains elusive. To address this, we intranasally inoculated mice with Streptococcus pneumoniae in the presence or absence of enzymes targeting pulmonary hyaluronan and heparan sulfate, followed by characterization of subsequent disease pathology, pulmonary inflammation, and lung barrier dysfunction. Enzymatic degradation of hyaluronan and heparan sulfate exacerbated pneumonia in mice, as evidenced by increased disease scores and alveolar neutrophil recruitment. However, targeting epithelial hyaluronan further exacerbated systemic disease, indicated by elevated splenic bacterial load and plasma levels of pro-inflammatory cytokines. In contrast, enzymatic cleavage of heparan sulfate resulted in increased bronchoalveolar bacterial burden, lung damage and pulmonary inflammation in mice infected with Streptococcus pneumoniae. Accordingly, heparinase-treated mice also exhibited disrupted lung barrier integrity as evidenced by higher alveolar edema scores and vascular protein leakage into the airways. This finding was corroborated in a human alveolus-on-a-chip platform, confirming that heparinase treatment also disrupts the human lung barrier. Notably, enzymatic pre-treatment with either hyaluronidase or heparinase also rendered human epithelial cells more sensitive to pneumococcal-induced barrier disruption, as determined by transepithelial electrical resistance measurements, consistent with our findings in murine pneumonia. Taken together, these findings demonstrate the importance of intact hyaluronan and heparan sulfate in controlling pneumococcal virulence, pulmonary inflammation, and epithelial barrier function.

molecular biology↗

bioRxiv2021

Inulin nanostructures: The sweet-spot of carbohydrate self-assembly

Supramolecular architecture formation by the self-assembly of proteins and nucleic acids is well studied. Yet, the spontaneous organization of oligosaccharides, the most common polymers in nature, is less explored. Here, using inulin as a model, we identify the "sweet spot" length for oligosaccharide assembly. Inulin forms discrete spheres in a concentration-dependent manner. Size-based fractionation displayed markedly different aggregation morphologies. Based on these findings, we believe that carbohydrates could become an important source for novel self-assembling materials.

biophysics↗

bioRxiv2021

Protein secondary structure affects glycan clustering in native mass spectrometry

Infection with human noroviruses (hNoV) for the vast majority of strains requires attachment of the viral capsid to histo blood group antigens (HBGA). The HBGA binding pocket is formed by dimers of the protruding domain (P dimers) of the capsid protein VP1. Several studies have focused on HBGA binding to P dimers, reporting binding affinities and stoichiometries. However, nuclear magnetic resonance spectroscopy (NMR) and native mass spectrometry (MS) analyses yielded incongruent dissociation constants (KD) for binding of HBGAs to P dimers and, in some cases, disagreed whether glycans bind at all. We hypothesized that glycan clustering during electrospray ionization in native MS critically depends on the physicochemical properties of the protein studied. It follows that the choice of the reference protein is crucial. We analyzed carbohydrate clustering using various P dimers and eight non-glycan binding proteins serving as possible references. Data from native and ion mobility MS indicate that the mass fraction of {beta}-sheet has a strong influence on the degree of glycan clustering. Therefore, the determination of specific glycan binding affinities from native MS must be interpreted cautiously.

biophysics↗

bioRxiv2019

High-throughput prediction of MHC Class I and Class II neoantigens with MHCnuggets

Computational prediction of binding between neoantigen peptides and major histocompatibility complex (MHC) proteins is an emerging biomarker for predicting patient response to cancer immunotherapy. Current neoantigen predictors focus on in silico estimation of MHC binding affinity and are limited by low positive predictive value for actual peptide presentation, inadequate support for rare MHC alleles and poor scalability to high-throughput data sets. To address these limitations, we developed MHCnuggets, a deep neural network method to predict peptide-MHC binding. MHCnuggets is the only method to handle binding prediction for common or rare alleles of MHC Class I or II, with a single neural network architecture. Using a long short-term memory network (LSTM), MHCnuggets accepts peptides of variable length and is capable of faster performance than other methods. When compared to methods that integrate binding affinity and HLAp data from mass spectrometry, MHCnuggets yields a fourfold increase in positive predictive value on independent MHC-bound peptide (HLAp) data. We applied MHCnuggets to 26 cancer types in TCGA, processing 26.3 million allele-peptide comparisons in under 2.3 hours, yielding 101,326 unique candidate immunogenic missense mutations (IMMs). Predicted-IMM hotspots occurred in 38 genes, including 24 driver genes. Predicted-IMM load was significantly associated with increased immune cell infiltration (p<2e-16) including CD8+ T cells. Notably, only 0.16% of predicted immunogenic missense mutations were observed in >2 patients, with 61.7% of these derived from driver mutations. Our results provide a new method for neoantigen prediction with high performance characteristics and demonstrate its utility in large data sets across human cancers.\n\nSynopsisWe developed a new in silico predictor of Major Histocompatibility Complex (MHC) ligand binding and demonstrated its utility to assess potential neoantigens and immunogenic missense mutations (IMMs) in 6613 TCGA patients.

immunology↗