Biology subjects

Levy, Y.

Publications and source records attributed to Levy, Y..

At least 19 recordsLinked to original sources

bioRxiv2026

Concurrent cognitive, behavioral and systemic physiological features mark preclinical Alzheimer's disease

Alzheimer's disease (AD) pathology accumulates years before overt cognitive impairment, yet its multidomain phenotype remains poorly defined. To limit confounding by uncontrolled comorbidities, we assembled a cohort of 644 older adults self-reporting intact cognition, free of systemic or neurological diseases and managed medically for risk factors. Plasma p-tau217 was elevated in 20%, and 93 of these were cognitively unimpaired. This biomarker-defined, risk factor -controlled preclinical AD subgroup showed subtle, concurrent cognitive and behavioral differences, accompanied by modest differences in physiological measures within clinical reference ranges, and alongside higher biomarkers of astrogliosis (GFAP) and neurodegeneration (NfL). Reported modifiable risk factor histories were not associated with p-tau217 when effectively managed. A proof-of-concept AI classifier integrating routine multidomain clinical data improved identification of p-tau217-high individuals relative to cognitive screening alone. These findings indicate preclinical AD is not phenotypically silent at the group level and support multidomain-based screening for biomarker testing and early-detection.

neuroscience↗

bioRxiv2026

Intrinsically disordered regions influence protein motif usage in Drosophila Ubx and AbdA Hox proteins

Hox proteins are evolutionary conserved transcription factors guiding morphogenesis in all bilaterian animals. Conservation mainly concerns the homeodomain (HD), a helix-turn-helix DNA binding motif and flanking sequences close by. Besides, sequences are poorly conserved and disordered. Here we investigate how poorly conserved Intrinsically Disordered Regions (IDRs) influence Hox protein activity, focusing on a closely related Drosophila Hox pair and using a wide range of in vivo function. Studying Drosophila Ultrabithorax (Ubx) and AbdominalA (AbdA) proteins, we find that conserved protein motifs close to the HD are differently used in Ubx and AbdA. Coarse-grained molecular dynamics simulations identify a large set of intra-molecular interactions, notably involving poorly conserved disordered regions upstream and downstream the HD. Swapping these regions in Ubx and AbdA alters protein functions and switches motif dependency in some biological situations. This suggests that IDRs, predicted to mediate large and dynamic sets of intramolecular interactions, influence protein motif usage in a context dependent manner.

developmental biology↗

bioRxiv2026

Comparison of Four Assays That Measure Antibodies to Ebola Virus Glycoprotein

The accurate measurement of Ebola virus (EBOV)-specific antibody responses is crucial to assessing immunity induced by EBOV infection or vaccination. For this purpose, the Filovirus Animal Nonclinical Group (FANG) anti-EBOV glycoprotein (GP1,2) ELISA is considered the "gold-standard". However, it has limitations such as high repeat-rates and variability, and low throughput. Here, we describe two new alternative assays: a Single-Molecule Assay Planar EBOV GP1,2 ELISA and a multiplexed EBOV GP1,2, EBOV nucleoprotein, and EBOV Viral Protein 40 Luminex assay, and compare these with two versions of the FANG ELISA. Samples were selected from participants receiving vaccine or placebo in a randomized, placebo-controlled, double-blinded study of two EBOV vaccines (PREVAIL 1), and a longitudinal cohort study of Ebola virus disease (EVD) survivors and their close contacts (PREVAIL 3). All four assays were concordant in their measurements of anti-EBOV GP1,2-specific immunoglobulin G responses, allowing for the determination of conversion equations for antibody measurements across assays. In addition, all four showed a similar ability to distinguish vaccine recipients from placebo recipients and EVD survivors from their close contacts. Compared to the FANG assays, the Quanterix and Luminex assays had lower variability, lower repeat rates, and higher throughput, making them good alternatives for future studies.

immunology↗

bioRxiv2026

Joint mechanistic modeling of viral and antibody responses to vaccines in non-human primates to quantify SARS-CoV-2 mechanistic correlates of protection

In a global SARS-CoV-2 landscape of hybrid immunity resulting from a vaccinated worldwide population and the emergence of new variants able to escape immunity, the identification and quantification of correlates of protection (CoPs) is crucial for the adaptation of next-generation SARS-CoV-2 vaccines. Antibodies and their neutralizing capacity have been identified as reliable mechanistic CoPs. Here, we proposed an original mechanistic model jointly describing viral and antibody dynamics, and their mutual interactions, observed after SARS-CoV-2 infection in non-human primates (NHPs) with distinct immunological backgrounds. From the model, the concentration of neutralizing antibodies being protective against viral infection spreading was derived using the reproduction number. Counterfactual simulations were also performed to better understand and validate immune mechanisms driving immune control. The model was estimated on viral, binding (bAb), and neutralizing (nAb) antibody dynamics, with data collected in 34 naive and convalescent NHPs. Animals were involved in a preclinical study evaluating two next-generation protein-based vaccines targeting the RBD of the Spike protein to CD40-expressing cells, and the original BNT162b2 mRNA vaccine against Delta SARS-CoV-2 infection. Our model validated the functionality of nAb to neutralize viruses as the primary mechanism of protection. An inhibitory antibody concentration against Delta variant of 20 AU/mL was deemed protective against Delta infection in naive animals. Moreover, we showed the strong benefit of hybrid immunity to induce faster and more protective antibody responses pointing out the crucial role of the memory B-cell immune response in viral control. Finally, an additional effect of natural immunity beyond antibodies and enhancing the elimination of infected cells was identified, suggesting the potential role of the T-cell response in viral control. Our model showed the benefit of combining information from both viral and antibody responses to better qualitatively and quantitatively inform on their protective capacity against Delta SARS-CoV-2 infection.

immunology↗

bioRxiv2025

Fluorescence lifetime-based biosensor for monitoring compartmentalized autophagy dynamics in the intact mammalian brain

Autophagy is a key process in regulation of neuronal development, plasticity, and local metabolism. Yet, autophagy dynamics and regulation within intact neuronal circuits remains poorly understood. Here, we developed a pH sensitive fluorescence lifetime-based imaging method which allows to monitor autophagy dynamics in the living mouse brain. This approach allowed us to uncover compartmentalized autophagic activity across soma, dendrites, and axons of layer 2/3 cortical neurons. We found pronounced differences in dendritic dynamics of autophagic vesicles in vivo, where distal dendrites showed elevated vesicle motility compared to proximal dendrites. Notably, sensory experience modulated dendritic autophagy dynamics in the somatosensory cortex. We further combined in vivo autophagy imaging with disease associated genetic perturbation to uncover novel autophagy related phenotypes. Altogether, this approach highlights the spatial and functional complexity of autophagy in the intact mammalian brain and establishes a framework for investigating its role in synaptic regulation during development, plasticity, and aging.

neuroscience↗

bioRxiv2025

Hair follicle stem cell fate supports distinct clinical endotypes in Hidradenitis Suppurativa

Hidradenitis suppurativa (HS) is a severe skin disorder affecting 1% of the global population, with a complex and poorly understood pathogenesis involving aberrant keratinization and autoinflammation. It remains unclear whether autoinflammatory events precede or follow hyperkeratotic changes in hair follicle (HF) epithelia. Using single-cell RNA sequencing, we characterized HF cell populations in HS patients and investigated their role in disease pathogenesis. We uncovered two distinct differentiation trajectories of HF stem cells (HF-SCs): one leading to interfollicular epidermis (IFE) basal cells enriched in inflammatory pathways, and another giving rise to outer root sheath (ORS) cells associated with keratinization. In HS lesions, both populations displayed altered inflammatory phenotypes and were closely linked to immune cell infiltration, pointing to a role in disease heterogeneity. By integrating clinical features with HF cell composition from 49 HS patients, we identified three major endotypes: (i) an inflammatory subtype, marked by T cell infiltration and an expansion of IFE basal cells; (ii) a keratinizing subtype, characterized by ORS enrichment and minimal inflammation; and a mixed subtype, exhibiting features of follicular remodeling, fistula formation, and variable immune involvement. These findings provide novel insights into the epithelial-immune interactions that drive HS and support a stratified therapeutic approach tailored to the specific HF dysfunctions of each patient subgroup. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=49 SRC="FIGDIR/small/656362v1_ufig1.gif" ALT="Figure 1"> View larger version (14K): org.highwire.dtl.DTLVardef@4e8af9org.highwire.dtl.DTLVardef@85f61borg.highwire.dtl.DTLVardef@e085a7org.highwire.dtl.DTLVardef@66d804_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2025

'What' and 'where' brain-wide pathways are dominated by internal strategies

It is long thought that higher-order sensory processing is divided into two specialized cortical streams that encode in parallel either the identity of an object or its location (i.e., what and where streams). Here, using the mouse whisker system, we challenge this concept by demonstrating an existence of two alternating brain-wide (beyond cortex) subnetworks that are not primarily driven by external parameters, but rather by internal strategies. We combine simultaneous brain-wide neuronal recordings in mice trained to identify or locate a certain stimulus. We find that mice deploy either an active search or a passive sensation strategy during task performance. These strategies respectively drive two distinct and brain-wide subnetworks, frontal and posterior, regardless of the type of task performed. The posterior subnetwork encoded additional internal strategic parameters such as trial history, the first task of the day, and training history. A subgroup of trials that were not dominated by frontal cortex, contained meaningful task information in the posterior cortex and several thalamic areas. Integrated together, these two subnetworks may comprise normal cognitive function.

neuroscience↗

bioRxiv2024

Ordered and disordered regions of the Origin Recognition Complex (ORC) combine to direct in-vivo binding at symmetric and non-symmetric motif sequences.

The Origin Recognition Complex (ORC) seeds replication-fork formation by binding to DNA replication origins, which in budding yeast contain a 17bp DNA motif. High resolution structure of the ORC-DNA complex revealed two base-interacting elements: a disordered basic patch (Orc1-BP4) and an insertion helix (Orc4-IH). To define the ORC elements guiding its DNA binding in-vivo, we mapped genomic locations of 38 designed ORC mutants, revealing that different ORC elements guide binding at different sites. At silencing-associated sites lacking the motif, ORC binding and activity were fully explained by a BAH domain. Within replication origins, we reveal two dominating motif variants showing differential binding modes and symmetry: an asymmetric motif whose binding requires Orc1-BP4 and Orc4-IH, and a symmetric one where another basic patch, Orc1-BP3, can replace Orc4-IH. Disordered basic patches are therefore key for ORC-motif binding in-vivo, and we discuss how these conserved, minor-groove interacting elements can guide specific ORC-DNA recognition.

cell biology↗

bioRxiv2023

Rapid long-distance migration of RPA on single stranded DNA occurs through intersegmental transfer utilizing multivalent interactions

Replication Protein A (RPA) is a single stranded DNA (ssDNA) binding protein that coordinates diverse DNA metabolic processes including DNA replication, repair, and recombination. RPA is a heterotrimeric protein with six functional oligosaccharide/oligonucleotide (OB) domains and flexible linkers. Flexibility enables RPA to adopt multiple configurations and is thought to modulate its function. Here, using single molecule confocal fluorescence microscopy combined with optical tweezers and coarse-grained molecular dynamics simulations, we investigated the diffusional migration of single RPA molecules on ssDNA under tension. The diffusion coefficient D is the highest (20,000 nucleotides2/s) at 3 pN tension and in 100 mM KCl and markedly decreases when tension or salt concentration increases. We attribute the tension effect to intersegmental transfer which is hindered by DNA stretching and the salt effect to an increase in binding site size and interaction energy of RPA-ssDNA. Our integrative study allowed us to estimate the size and frequency of intersegmental transfer events that occur through transient bridging of distant sites on DNA by multiple binding sites on RPA. Interestingly, deletion of RPA trimeric core still allowed significant ssDNA binding although the reduced contact area made RPA 15-fold more mobile. Finally, we characterized the effect of RPA crowding on RPA migration. These findings reveal how the high affinity RPA-ssDNA interactions are remodeled to yield access, a key step in several DNA metabolic processes. SignificanceReplication Protein A (RPA) binds to the exposed single stranded DNA (ssDNA) during DNA metabolism. RPA dynamics are essential to reposition RPA on ssDNA and recruit downstream proteins at the bound site. Here in this work, we perform a detailed biophysical study on dynamics of yeast RPA on ssDNA. We show that RPA can diffuse on ssDNA and is affected by tension and salt. Our observations are best explained by the intersegmental transfer model where RPA can transiently bridge two distant DNA segments for its migration over long distances. We further dissect the contributions of the trimerization core of RPA and other adjacent RPA molecules on RPA migration. This study provides detailed experimental and computational insights into RPA dynamics on ssDNA.

biophysics↗

bioRxiv2023

Sound-mediated nucleation and growth of amyloid fibrils

Mechanical energy, specifically in the form of ultrasound, can induce pressure variations and temperature fluctuations when applied to an aqueous media. These conditions can both positively and negatively affect protein complexes, consequently altering their stability, folding patterns, and self-assembling behavior. Despite much scientific progress, our current understanding of the effects of ultrasound on the self-assembly of amyloidogenic proteins remains limited. In the present study, we demonstrate that when the amplitude of the delivered ultrasonic energy is sufficiently low, it can induce refolding of specific motifs in protein monomers, which is sufficient for primary nucleation; this has been revealed by MD. These ultrasound-induced structural changes are initiated by pressure perturbations and are accelerated by a temperature factor. Furthermore, the prolonged action of low-amplitude ultrasound enables the elongation of amyloid protein nanofibrils directly from natively folded monomeric lysozyme protein, in a controlled manner, until it reaches a critical length. Using solution X-ray scattering, we determined that nanofibrillar assemblies, formed either under the action of sound or from natively fibrillated lysozyme, share identical structural characteristics. Thus, these results provide insights into the effects of ultrasound on fibrillar protein self-assembly and lay the foundation for the potential use of sound energy in protein chemistry. Significance StatementUnderstanding how and why proteins form amyloid fibrils is crucial for research into various diseases, including neurodegeneration. Ultrasound is routinely used in research settings as a tool for generating amyloid seeds (nucleation sites) from mature fibrils, which accelerate the rate of fibril growth. However, ultrasound can have various effects on aqueous media including temperature, extreme shear, and free radicals. Here we show that when the ultrasound parameters are precisely adjusted, they can be utilized as a tool for amyloid growth directly from the natively folded monomers. Thus, it is possible to induce minor changes in the folding of proteins, which trigger nucleation and accelerate amyloid growth. This knowledge lays the foundation for the potential use of sound in protein chemistry.

biochemistry↗

bioRxiv2023

Muscle fibre size and myonuclear positioning in trained and aged humans

Myonuclear domain (MND) is the theoretical volume of cytoplasm within which a myonucleus is responsible for transcribing DNA. Changes in myonuclear number, organisation, and myonuclear domain size are associated with exercise adaptations and ageing. However, data on satellite cell activation, changes in MND volumes and myonuclear arrangement following exercise are inconsistent. Additionally, whether MNDs and myonuclear arrangement are altered with age remains unclear. The aim of the present investigation was therefore to investigate relationships between age and activity status and myonuclear numbers and organisation. Muscle fibres from younger trained (YT) and older trained (OT) individuals were compared with age-matched untrained counterparts (YU and OU). Serial, optical z-slices were acquired throughout isolated muscle fibres and analysed to give 3D coordinates for myonuclei and muscle fibre dimensions, respectively. As expected, mean cross-sectional area (CSA) (m2) of muscle fibres from OU was 29-42% smaller compared to the other groups. Number of nuclei relative to fibre CSA was 87% greater in OU compared to YU muscle fibres (P < 0.05). Additionally, scaling of myonuclear domain volume with fibre size was altered in older untrained individuals. Myonuclear arrangement, on the other hand, was similar across groups. These data indicate that regular endurance exercise throughout the lifespan may preserve the size of single muscle fibres in older age and maintain the relationship between fibre size and MND volumes. Inactivity, however, may result in reduced muscle fibre size and disrupted relationship between fibre size and MND volumes. Plain Language SummaryIn this study, we examined the relationship between physical activity and the characteristics of muscle fibres in individuals of different age groups. We focused on a concept called the myonuclear domain (MND), which refers to the volume surrounding muscle nuclei or myonuclei that house the genome. We wanted to understand how changes in myonuclear number, organisation, and MND size were influenced by exercise and aging. To do this, we compared muscle fibres from younger trained individuals, older trained individuals, and age-matched untrained individuals. The results showed that the average size of muscle fibres in the untrained older individuals was smaller compared to the other groups. Moreover, the number of nuclei relative to fibre size was significantly higher in the untrained older individuals. However, myonuclear arrangement was similar across all groups. These findings suggest that regular endurance exercise throughout life may help maintain muscle fibre size, myonuclear numbers, MND volumes, and myonuclear organisation in older individuals. Conversely, inactivity can lead to reduced muscle fibre size and disrupted relationship between fibre size and MND volumes. These results have important implications for understanding the effects of exercise and aging on muscle health.

physiology↗

bioRxiv2023

ACE2 mimetic antibody potently neutralizes all SARS-CoV-2 variants and fully protects in XBB.1.5 challenged monkeys

The rapid evolution of SARS-CoV-2 to variants with improved transmission efficiency and reduced sensitivity to vaccine-induced humoral immunity has abolished the protective effect of licensed therapeutic human monoclonal antibodies (mAbs). To fill this unmet medical need and protect vulnerable patient populations, we isolated the P4J15 mAb from a previously infected, vaccinated donor, with <20 ng/ml neutralizing activity against all Omicron variants including the latest XBB.2.3 and EG.1 sub-lineages. Structural studies of P4J15 in complex with Omicron XBB.1 Spike show that the P4J15 epitope shares [~]93% of its buried surface area with the ACE2 contact region, consistent with an ACE2 mimetic antibody. Although SARS-CoV-2 mutants escaping neutralization by P4J15 were selected in vitro, these displayed lower infectivity, poor binding to ACE2, and the corresponding escape mutations are accordingly rare in public sequence databases. Using a SARS-CoV-2 XBB.1.5 monkey challenge model, we show that P4J15 confers complete prophylactic protection. We conclude that the P4J15 mAb has potential as a broad-spectrum anti-SARS-CoV-2 drug.

microbiology↗

bioRxiv2023

Allochthonous groundwater microorganisms affect coastal seawater microbial abundance, activity and diversity

Submarine groundwater discharge (SGD) is a globally important process supplying nutrients and trace elements to the coastal environment, thus playing a pivotal role in sustaining marine primary productivity. Along with nutrients, groundwater also contains allochthonous microbes that are discharged from the terrestrial subsurface into the sea. Currently, little is known about the interactions between groundwater-borne and coastal seawater microbial populations, and their role upon introduction to coastal seawater populations. Here, we investigated seawater microbial abundance, activity and diversity in a site strongly influenced by SGD (in-situ observations), and through laboratory-controlled bottle incubations mimicking different mixing scenarios between SGD (either ambient or filtered through 0.1 {micro}m/0.22 {micro}m) and seawater. Our results demonstrate that the addition of <0.1 {micro}m SGD stimulated heterotrophic activity and increased microbial abundance compared to control, whereas <0.22 {micro}m filtration treatments induced primary productivity rates and Synechococcus growth. Amplicon sequencing of the 16S rRNA gene showed a strong shift from a SAR11-rich community in the reference SGD-unaffected coastal samples to a Rhodobacteraceae-dominated one in the <0.1 {micro}m treatment, in agreement with their in-situ enrichment in the SGD field site. These results suggest that despite the significant nutrient input, microbes delivered by SGD may affect the abundance, activity and diversity of intrinsic microbes in coastal seawater. Our results highlight the cryptic interplay between groundwater and seawater microbes in coastal environments, which has important implications for carbon cycling and climate. Key PointsO_LIGroundwater discharge into the coastal zone delivers both nutrients and allochthonous microbes. C_LIO_LIGroundwater microbes interact with seawater populations, by which affecting the delicate autotroph-heterotroph balance. C_LIO_LISubterranean microbial processes are key drivers of food webs, potentially affecting biogenic carbon fluxes in the ocean and climate. C_LI

microbiology↗

bioRxiv2023

Functional Ambidexterity of an Ancient Nucleic Acid-Binding Domain

Homochirality of biopolymers emerged early in the history of life on Earth, nearly 4 billion years ago. Whether the establishment of homochirality was the result of abiotic physical and chemical processes, or biological selection, remains unknown. However, given that significant events in protein evolution predate the last universal common ancestor, the history of homochirality may have been written into some of the oldest protein folds. To test this hypothesis, the evolutionary trajectory of the ancient and ubiquitous helix-hairpin-helix (HhH) protein family was analyzed for functional robustness to total chiral inversion of just one binding partner. Against expectations, functional ambidexterity was observed across the entire trajectory, from phase separation of HhH peptides with RNA to dsDNA binding of the duplicated (HhH)2-Fold. Moreover, dissociation kinetics, mutational analysis, and molecular dynamics simulations revealed significant overlap between the binding modes of a natural and a mirror-image protein to natural dsDNA. These data suggest that the veil between worlds with alternative chiral preferences may not be as impenetrable as is often assumed, and that the HhH protein family is an intriguing exception to the dogma of reciprocal chiral substrate specificity proposed by Milton and Kent (Milton et al. Science 1992).

biophysics↗

bioRxiv2022

Exercise induces myonuclear remodelling in humans independently of age

Age-related decline in skeletal muscle structure and function can be mitigated by regular exercise. However, the precise mechanisms that govern this are not fully understood. The nucleus plays an active role in translating forces into biochemical signals (mechanotransduction), with nuclear lamina protein Lamin A regulating nuclear shape, nuclear mechanics, and ultimately gene expression. Defective Lamin A expression causes muscle pathologies and premature ageing syndromes, but the roles of nuclear structure and function in physiological ageing and in exercise adaptations remain obscure. Here, we isolated single muscle fibres and carried out detailed morphological and functional analyses on myonuclei from young and older exercise-trained individuals. Strikingly, myonuclei from trained individuals were more spherical, less deformable, and contained a thicker nuclear lamina than untrained individuals. Complementary to this, exercise resulted in increased levels of Lamin A and increased myonuclear stiffness in mice. We conclude that exercise is associated with myonuclear remodelling, independently of age, which may contribute to the preservative effects of exercise on muscle function throughout the lifespan. Key pointsO_LIThe nucleus plays an active role in translating forces into biochemical signals C_LIO_LIMyonuclear aberrations in a group of muscular dystrophies called laminopathies suggest that the shape and mechanical properties of myonuclei are important for maintaining muscle function. C_LIO_LIHere, we present striking differences in myonuclear shape and mechanics associated with exercise, in both young and old humans. C_LIO_LIMyonuclei from trained individuals were more spherical, less deformable, and contained a thicker nuclear lamina than untrained individuals. C_LIO_LIWe conclude that exercise is associated with age-independent myonuclear remodelling, which may help to maintain muscle function throughout the lifespan. C_LI

cell biology↗

bioRxiv2022

An Effective mRNA-LNP Vaccine Against the Lethal Plague Bacterium

Plague is a rapidly deteriorating contagious disease that has killed millions of people during the history of mankind and is caused by the gram-negative bacterium Yersinia pestis. Currently, the disease is treated effectively with antibiotics. However, in the case of an outbreak caused by a multiple-antibiotic-resistant strain, alternative countermeasures are required. Despite the many efforts to develop a safe vaccine against the disease, there is still no vaccine approved for use in western countries. mRNA Lipid Nanoparticle (mRNA-LNP) vaccines have been demonstrated during the Covid-19 pandemic to be a versatile, clinically relevant, and rapidly manufactured vaccine platform. However, harnessing this platform for bacterial pathogens remains a formidable challenge. Here, we describe the design of several mRNA-LNP vaccine versions against Y. pestis, based on the F1 capsular antigen. We demonstrate that mRNA-LNP vaccines encoding the F1 antigen with either no signal sequences or conjugated to human Fc, provide substantial cellular and humoral responses. Most importantly, these vaccine candidates fully protect animals against Y. pestis infection. The results of this study suggest that mRNA-LNPs can be effective as anti-bacterial vaccines, and further developed to combat other bacterial pathogens, which are urgently needed, given the looming threat of antibiotic resistance. One-Sentence SummaryA novel mRNA-LNP vaccine against Y. pestis, the etiological agent of plague and the first documented mRNA-LNP vaccine to protect against a lethal bacterial pathogen infection.

microbiology↗

bioRxiv2022

Peptide-RNA Coacervates as a Cradle for the Evolution of Folded Domains

Peptide-RNA coacervates can result in the concentration and compartmentalization of simple biopolymers. Given their primordial relevance, peptide-RNA coacervates may have also been a key site of early protein evolution. However, the extent to which such coacervates might promote or suppress the exploration of novel peptide conformations is fundamentally unknown. To this end, we used electron paramagnetic resonance (EPR) spectroscopy to characterize the structure and dynamics of an ancient and ubiquitous nucleic acid binding element, the helix-hairpin-helix (HhH) motif, alone and in the presence of RNA, with which it forms coacervates. Double electron-electron resonance (DEER) spectroscopy applied to singly labeled peptides containing one HhH motif reveals the presence of dimers, even in the absence of RNA, and transient -helical character. Moreover, dimer formation is promoted upon RNA binding and was detectable within peptide-RNA coacervates. The distance distributions between spin labels are consistent with the symmetric (HhH)2-Fold, which is generated upon duplication and fusion of a single HhH motif and traditionally associated with dsDNA binding. These results support the hypothesis that coacervates are a unique testing ground for peptide oligomerization and that phase-separating peptides could have been a resource for the construction of complex protein structures via common evolutionary processes, such as duplication and fusion.

biophysics↗

bioRxiv2022

SARS-CoV-2 Omicron potently neutralized by a novel antibody with unique Spike binding properties

The SARS-CoV-2 Omicron variant exhibits very high levels of transmission, pronounced resistance to authorized therapeutic human monoclonal antibodies and reduced sensitivity to vaccine-induced immunity. Here we describe P2G3, a human monoclonal antibody (mAb) isolated from a previously infected and vaccinated donor, which displays picomolar-range neutralizing activity against Omicron BA.1, BA.1.1, BA.2 and all other current variants, and is thus markedly more potent than all authorized or clinically advanced anti-SARS-CoV-2 mAbs. Structural characterization of P2G3 Fab in complex with the Omicron Spike demonstrates unique binding properties to both down and up spike trimer conformations at an epitope that partially overlaps with the receptor-binding domain (RBD), yet is distinct from those bound by all other characterized mAbs. This distinct epitope and angle of attack allows P2G3 to overcome all the Omicron mutations abolishing or impairing neutralization by other anti-SARS-COV-2 mAbs, and P2G3 accordingly confers complete prophylactic protection in the SARS-CoV-2 Omicron monkey challenge model. Finally, although we could isolate in vitro SARS-CoV2 mutants escaping neutralization by P2G3 or by P5C3, a previously described broadly active Class 1 mAb, we found these viruses to be lowly infectious and their key mutations extremely rare in the wild, and we could demonstrate that P2G3/P5C3 efficiently cross-neutralized one anothers escapees. We conclude that this combination of mAbs has great prospects in both the prophylactic and therapeutic settings to protect from Omicron and other VOCs.

immunology↗