Biology subjects

Jovine, L.

Publications and source records attributed to Jovine, L..

9 recordsLinked to original sources

bioRxiv2026

How a single sperm protein recognizes highly divergent egg coat components

For successful fertilization, sperm must recognize and penetrate the egg coat, called vitelline envelope (VE) or, in mammals, zona pellucida (ZP). In abalone, sperm acrosomal protein lysin dissolves the VE by binding the N-terminal region of egg coat subunit VERL, encompassing 22 ZP-N domains (VR1-22), but how sperm engages other VE components remains unclear. Here we report free and lysin-bound structures of VEZP14-N1, the single ZP-N repeat of VERL paralog VEZP14, together with biochemical and biophysical data. Despite [~]20% sequence identity to VRs, VEZP14-N1 presents a structurally equivalent surface to lysin, indicating that conserved interaction geometry underlies recognition. Binding is asymmetric: lysin is conformationally rigid, whereas VEZP14-N1 requires homodimer dissociation and induced-fit rearrangements to bind -- kinetic costs that limit affinity to an intermediate level, consistent with a decoy role sequestering lysin from VERL. These findings illuminate how one sperm protein binds divergent egg coat targets, with implications for ZP-N repeat function in vertebrate gamete recognition.

biophysics↗

bioRxiv2025

Uromodulin velcro sheets and their interaction with uropathogenic E. coli

Homopolymeric uromodulin (UMOD), the most abundant protein in human urine, protects against urinary tract infections (UTI) by acting as a decoy for uropathogenic E. coli (UPEC). Using cryo-electron tomography (ET), we show that urinary UMOD filaments naturally form sheets that interact with UPEC type I pili. Sheet formation is salt-dependent, and we resolve their high-resolution structure using single-particle cryo-electron microscopy (EM). This reveals a lateral interface between polymers, whose mutation disrupts UMOD filament bundle formation in mammalian cells. Branchless egg coat protein ZPD also forms sheets, and cryo-ET of elastase-treated UMOD indicates that absence of N-terminal branches promotes the stacking of UMOD-like protein sheets into thick 3D matrices. These results rationalize early observations of salt-dependent aggregation of UMOD, explain how UMOD can assemble into extended velcro-like structures that efficiently inactivate a multitude of adhesive UPEC pili, and suggest how UMOD-like molecules can generally organize into supramolecular structures of variable thickness.

biophysics↗

bioRxiv2025

Structural basis of ZP2-targeted immunocontraception

Monoclonal antibody IE-3 prevents mouse fertilization by binding ZP2, a major component of the oocyte-specific zona pellucida (ZP). We show that an IE3-derived single-chain variable fragment (scFV) is sufficient for blocking fertilization in vitro and determine the structural basis of IE-3/ZP2 recognition. The high-affinity of this interaction depends on induced fit of the epitope, offering insights for non-hormonal immunocontraceptive design without off-target effects.

biophysics↗

bioRxiv2023

Deep learning insights into the architecture of the mammalian egg-sperm fusion synapse

A crucial event in sexual reproduction is when haploid sperm and egg fuse to form a new diploid organism at fertilization. In mammals, direct interaction between egg JUNO and sperm IZUMO1 mediates gamete membrane adhesion, yet their role in fusion remains enigmatic. We used AlphaFold to predict the structure of other extracellular proteins essential for fertilization to determine if they could form a complex that may mediate fusion. We first identified TMEM81, whose gene is expressed by mouse and human spermatids, as a protein having structural homologies with both IZUMO1 and another sperm molecule essential for gamete fusion, SPACA6. Using a set of proteins known to be important for fertilization and TMEM81, we then systematically searched for predicted binary interactions using an unguided approach and identified a pentameric complex involving sperm IZUMO1, SPACA6, TMEM81 and egg JUNO, CD9. This complex is structurally consistent with both the expected topology on opposing gamete membranes and the location of predicted N-glycans not modeled by AlphaFold-Multimer, suggesting that its components could organize into a synapse-like assembly at the point of fusion. Finally, the structural modeling approach described here could be more generally useful to gain insights into transient protein complexes difficult to detect experimentally. Impact statementStructural modeling with AlphaFold-Multimer was used to investigate extracellular protein interactions involved in mammalian egg-sperm recognition, suggesting a putative pentameric complex that includes TMEM81, a sperm protein not previously involved in gamete recognition.

biophysics↗

bioRxiv2023

Architecture of the Vertebrate Egg Coat and Structural Basis of the ZP2 Block to Polyspermy

Post-fertilization cleavage of glycoprotein ZP2, a major subunit of egg zona pellucida (ZP) filaments, is crucial for mammalian reproduction by irreversibly blocking polyspermy. ZP2 processing is thought to inactivate a sperm-binding activity located upstream of the proteins cleavage site; however, its molecular consequences and connection with ZP hardening are unknown. Here we report X-ray crystallographic, cryo-EM and biochemical studies showing that cleavage of ZP2 triggers its oligomerization. Deletion of the ZP-N1 domain that precedes the cleavage site of mouse ZP2 allows it to homodimerize even without processing, and animals homozygous for this variant are subfertile by having a semi-hardened ZP that allows sperm attachment but hinders penetration. Combined with the structure of a native egg coat filament, which reveals the molecular basis of heteromeric ZP subunit interaction, this suggests that oligomerization of cleaved ZP2 cross-links the ZP, rigidifying it and making it physically impenetrable to sperm.

biophysics↗

bioRxiv2022

Complementing machine learning-based structure predictions with native mass spectrometry

The advent of machine learning-based structure prediction algorithms such as AlphaFold2 (AF2) has moved the generation of accurate structural models for the entire cellular protein machinery into the reach of the scientific community. However, structure predictions of protein complexes are based on user-provided input and may therefore require experimental validation. Mass spectrometry (MS) is a versatile, time-effective tool that provides information on post-translational modifications, ligand interactions, conformational changes, and higher-order oligomerization. Using three protein systems, we show that native MS experiments can uncover structural features of ligand interactions, homology models, and point mutations, that are undetectable by AF2 alone. We conclude that machine learning can be complemented with MS to yield more accurate structural models on the small and the large scale.

biophysics↗

bioRxiv2021

Archaeal origins of gamete fusion

Sexual reproduction consists of genome reduction by meiosis and subsequent gamete fusion. Presence of meiotic genes in prokaryotes suggests that DNA repair mechanisms evolved toward meiotic recombination; however, fusogenic proteins resembling those found in eukaryotes were not identified in prokaryotes. Here, we identify archaeal proteins that are homologs of fusexins, a superfamily of fusogens that mediate eukaryotic gamete and somatic cell fusion, as well as virus entry. The crystal structure of a trimeric archaeal Fusexin1 reveals novel features such as a six-helix bundle and an additional globular domain. Ectopically expressed Fusexin1 can fuse mammalian cells, and this process involves the additional domain and a conserved fusion loop. Archaeal fusexin genes exist within integrated mobile elements, potentially linking ancient archaeal gene exchanges and eukaryotic sex. One-Sentence SummaryCell membrane fusion proteins of viruses and eukaryotes are also present in archaea.

evolutionary biology↗

bioRxiv2020

Cryo-EM Structure of Native Human Uromodulin, a Zona Pellucida Module Polymer

Assembly of extracellular filaments and matrices mediating fundamental biological processes such as morphogenesis, hearing, fertilization and antibacterial defense is driven by a ubiquitous polymerization module known as zona pellucida (ZP) "domain". Despite the conservation of this element from hydra to human, no information is available on the filamentous conformation of any ZP module protein. Here we report the cryo-electron microscopy structure of uromodulin (UMOD)/Tamm-Horsfall protein, the most abundant protein in human urine and an archetypal ZP module-containing molecule, in its mature homopolymeric state. UMOD forms a one-start helix with an unprecedented 180-degree twist between subunits enfolded by interdomain linkers that have completely reorganized as a result of propeptide dissociation. Lateral interaction between filaments in the urine generates sheets exposing a checkerboard of binding sites to capture uropathogenic bacteria, and UMOD-based models of mammalian and avian heteromeric egg coat filaments identify a common sperm-binding region at the interface between subunits.

biophysics↗

bioRxiv2019

Molecular basis of egg coat cross-linking sheds light on ZP1-associated female infertility

Interaction between sperm and the egg zona pellucida (ZP) is the first step of mammalian fertilization, and ZP component ZP1 is important for fertility by covalently cross-linking ZP filaments into a matrix. Like ZP4, a structurally-related subunit absent in the mouse, ZP1 is predicted to contain an N-terminal ZP-N domain of unknown function. Characterization of ZP1 proteins carrying mutations from infertile patients suggests that, unlike in the mouse, filament cross-linking by ZP1 is crucial for human ZP assembly. We map the function of ZP1 to its ZP-N1 domain and determine crystal structures of ZP-N1 homodimers from a chicken homolog of ZP1. These reveal that ZP filament cross-linking is highly plastic and can be modulated by ZP1 fucosylation and, potentially, zinc sparks. Moreover, we show that ZP4 ZP-N1 forms non-covalent homodimers in chicken but not human. Together, these data identify human ZP1 cross-links as a promising target for non-hormonal contraception.

biophysics↗