Schmitt, M. P. | bioRxiv Science

bioRxiv2026

SMC Motor Proteins Operate at the Near-Minimal Forces for DNA Loop Extrusion

Loop extrusion by structural maintenance of chromosomes (SMC) complexes is essential for genome organization, yet the forces driving this process remain poorly understood. We present a coarse-grained model enabling predictive simulations of in vitro loop extrusion experiments at experimentally relevant time and length scales by matching parameters with concrete experiments. Using this model, we demonstrate that the extrusion forces generated by SMC motor proteins are just sufficient to overcome initial entropic barriers and sustain loop extrusion, highlighting that motors operate in the thermal regime. By measuring stalling tension directly, we confirm that they can be reliably determined by the Marko-Siggia equation and that varying grafting distances in experimental setups has only a marginal effect on the resulting tension. These results provide a predictive computation method for dissecting the mechanics of SMC driven genome folding.

biophysics↗

bioRxiv2026

Native Mass Spectrometry-Based Proteomics Reveals the Mechanism of Hemophore Release by Pathogenic Corynebacterium diphtheriae

Iron is an essential micronutrient for nearly all forms of life, including pathogenic microbes that must acquire it from their host during infection. At the host-pathogen interface, humans restrict microbial access to iron through nutritional immunity, which many pathogens overcome by secreting hemophores that scavenge extracellular heme (iron protoporphyrin IX). However, identifying hemophores and other ligand-binding proteins in complex proteomes remains challenging using conventional peptide-based bottom-up mass spectrometry (MS). Here, we introduce ProteoMIX (Proteome Analysis by Mixing), a function-based native top-down proteomics workflow that combines slow-mixing mode native MS with charge reduction to identify ligand-binding proteins directly from complex mixtures. Applying ProteoMIX to the Corynebacterium diphtheriae exoproteome identified ChtA30-314, an abundant soluble hemophore generated by proteolytic processing of the surface-exposed ChtA heme receptor. Using native top-down MS sequencing, cell fractionation, and gene deletion, we show that the protease DIP2069 releases ChtA30-314 by removing ChtAs transmembrane helix, producing a soluble proteoform that delivers heme to support microbial growth. In contrast, the related paralog ChtC is not processed, enabling C. diphtheriae to generate localization-specific heme-binding proteoforms from related gene products. Extending this approach to Staphylococcus aureus, ProteoMIX also revealed soluble IsdA hemophores that coexist with surface-anchored variants, demonstrating the generality of the method and suggesting that protease-mediated hemophore release may operate across gram-positive pathogens.

microbiology↗