Biology subjects

Vavra, J.

Publications and source records attributed to Vavra, J..

3 recordsLinked to original sources

bioRxiv2026

Resolving Heterogeneous Mechanical Domains via Physics-Aware Deep Clustering of Single-Molecule Force Spectroscopy Data

Many biological processes rely on mechanical forces, with protein molecules acting as key mediators. Understanding how proteins respond to mechanical stress is essential for conditions including cardiomyopathy and muscular dystrophy. Natural proteins such as dystrophin and utrophin are composed of heterogeneous folding domains with distinct mechanical properties; deciphering domain-level behavior provides insights into disease mechanisms and informs therapeutic strategies. Single-molecule force spectroscopy (SMFS) enables probing the mechanical properties of entire proteins, yet current approaches struggle to identify heterogeneous folding domains, particularly without prior knowledge. Here, we present the first automated framework to identify heterogeneous folding domains in SMFS data, applying both existing clustering methods and a novel physics-aware deep clustering architecture, LatentUnfold. LatentUnfold learns complementary latent representations from force magnitude and the force-extension physical relationship through dual autoencoders, jointly optimized for clustering assignments. We apply our framework to experimental SMFS data collected from a synthetic two-domain protein (ddFLN4-Titin I27) as well as natural protein constructs of dystrophin and utrophin, with Monte Carlo simulated datasets serving as controlled validation. For the synthetic protein, we recover mechanical properties consistent with previously reported values for each domain. For the natural proteins, we uncover two mechanically distinct domain populations - corresponding to the N-terminal domain and spectrin-like repeats - with differences in both unfolding force and contour length increase, and reveal different unfolding order between them for the first time. This work enables domain-level biological inference, overcoming prior limitations that relied on averaging and overlooked heterogeneity, thus advancing the understanding of mechanical behavior in protein unfolding.

biophysics↗

bioRxiv2026

Reconstructing 50 million years of Xenopus borealis evolution: three temporal strata of DNA rearrangements and persistent sex chromosome homomorphism

Genomic rearrangements are fundamental drivers of biodiversity, yet dynamics of structural evolution following polyploidization remain poorly understood. Genus Xenopus provides a valuable tool to study these phenomena. Utilizing the diploid X. tropicalis as a reference, we employed cytogenetic and genomic mapping to track the structural evolution of the allotetraploids X. borealis and X. laevis across a 50-million-year timeline. Based on chromosome morphometrics and C-banding patterns, we characterized the X. borealis pseudotetraploid karyotype (2n = 4x = 36), localizing the nucleolus organizer region (NOR) to chromosome 5L, U1 and U2 small nuclear DNAs to 1S and 8L, and 5S rDNA to nearly all chromosomes. Our analysis revealed 17 genomic rearrangements distributed within three temporal strata: ancestral (50-35 Mya), intermediate (35-15 Mya), and recent (< 15 Mya). Although we categorized chromosome 9/10 fusion as an ancestral rearrangement, the 2/9 translocation previously identified in X. mellotropicalis was absent in both studied allotetraploids. Furthermore, we tested for sex-specific structural polymorphism on the X. borealis W chromosome. Despite a large region of recombination suppression between the W and Z, no inversions were detected, indicating persistent sex chromosome homomorphism. Results are consistent with the expectation that tandem repeats such as NORs follow an asymmetric trajectory driven by a jumping mechanism and biased deletion, whereas small nuclear DNA loci are governed by copy number reduction-expansion dynamics. These findings indicate that structural rearrangements in Xenopus were not limited to punctuated bursts immediately following whole-genome duplication; rather, they accumulated over a prolonged evolutionary history, affecting the entire polyploid complement.

evolutionary biology↗

bioRxiv2024

Two operational modes of atomic force microscopy reveal similar mechanical properties for homologous regions of dystrophin and utrophin

Duchenne muscular dystrophy (DMD) is a lethal muscle disease caused by the absence of the protein dystrophin. Dystrophin is hypothesized to work as a molecular shock absorber that limits myofiber membrane damage when undergoing reversible unfolding upon muscle stretching and contraction. Utrophin is a dystrophin homologue that is under investigation as a protein replacement therapy for DMD. However, it remains uncertain whether utrophin can mechanically substitute for dystrophin. Here, we compared the mechanical properties of homologous utrophin and dystrophin fragments encoding the N terminus through spectrin repeat 3 (UtrN-R3, DysN-R3) using two operational modes of atomic force microscopy (AFM), constant speed and constant force. Our comprehensive data, including the statistics of force magnitude at which the folded domains unfold in constant speed mode and the time of unfolding statistics in constant force mode, show consistent results. We recover parameters of the energy landscape of the domains and conducted Monte Carlo simulations which corroborate the conclusions drawn from experimental data. Our results confirm that UtrN-R3 expressed in bacteria exhibits significantly lower mechanical stiffness compared to insect UtrN-R3, while the mechanical stiffness of the homologous region of dystrophin (DysN-R3) is intermediate between bacterial and insect UtrN-R3, showing greater similarity to bacterial UtrN-R3. SignificanceDuchenne muscular dystrophy (DMD) is a severe muscle wasting disorder caused by mutations in DMD gene encoding dystrophin. Utrophin, a fetal homologue of dystrophin, is under active investigation as a dystrophin replacement therapy for DMD. However, it is still unknown if it can substitute dystrophin mechanically. Here, we report mechanical properties of both utrophin and dystrophin fragments encoding the N terminus through spectrin repeat 3 (UtrN-R3, DysN-R3) using atomic force microscope (AFM) through two operational modes, constant speed and constant force. Our data, consistent across both modes, confirm that UtrN-R3 expressed in bacteria exhibits significantly lower mechanical stiffness than insect UtrN-R3. Additionally, bacterial DysN-R3 lies between bacterial and insect UtrN-R3 in terms of mechanical properties, leaning closer to bacterial UtrN-R3.

biophysics↗