Biology subjects

Vetrano, A.

Publications and source records attributed to Vetrano, A..

3 recordsLinked to original sources

bioRxiv2026

Structural studies of the C-terminal cysteine-rich domain of Aedes albopictus vitellogenin reveal an evolutionarily divergent, monomeric C-terminal cysteine knot (CTCK) architecture

Vitellogenins are essential transport lipoproteins and precursors to egg-yolk proteins in oviparous species. Several molecular structure studies have elucidated most of their multi-domain organization, yet the structure and function of their C-terminal cysteine knot (CTCK) domain have remained largely elusive. In this study, we present the 1.2 [A] resolution crystal structure of the recombinant CTCK domain from a vitellogenin isoform of the mosquito Aedes albopictus (Vg-CTCK). The molecular architecture reveals a CTCK fold defined by two antiparallel beta sheets stabilized by three intramolecular disulfide bonds, featuring a unique 12-amino acid insertion shaping an alpha helix positioned between the two main beta sheets. Analysis of the crystal packing and biophysical characterization in solution consistently confirms that recombinant Vg-CTCK is monomeric. To validate these findings in a native context, we employed ex vivo cross-linking mass spectrometry (XL-MS) on intact mosquito ovaries, which corroborated the molecular architecture of the Vg-CTCK observed in the crystal structure and highlighted the absence of inter-molecular cross-links. Collectively, our data highlight an evolutionarily divergent, monomeric assembly for the mosquito Vg-CTCK domain, challenging previous hypotheses that suggested this domain might facilitate vitellogenin oligomerization.

molecular biology↗

bioRxiv2025

Enantiopurity-Dependent Peptide Coacervates and Asymmetric Organocatalysis

Membraneless compartmentalization via liquid-liquid phase separation (LLPS) has emerged as a powerful strategy to organize biochemical reactions. Recently, peptide-based coacervates demonstrated the potential to function as microreactors by enhancing reaction kinetics through increased local concentrations and altered microenvironments. Here, we introduce an O-methylated diphenylalanine-based tripeptide LLLPFF-OCH3 containing an N-terminal proline, designed to undergo LLPS, and simultaneously function as an enantioselective organocatalyst. Comprehensive characterization via confocal microscopy, fluorescence recovery after photobleaching (FRAP), micro-Raman and attenuated total reflection infrared (ATR-IR) spectroscopy, diffusion-surface plasmon resonance (D-SPR), and molecular dynamics (MD) simulations revealed the formation of stable liquid droplets. In contrast, a racemic mixture of LLLPFF-OCH3 and DDDPFF-OCH3 failed to form liquid droplets and instead formed a solid precipitate, unveiling a critical role of enantiopurity in LLPS. Proof-of-concept catalytic studies proved enantioselective organocatalytic activity of the LLLPFF-OCH3 liquid coacervates. Beyond catalysis these results may have broader implications in understanding prebiotic chemistry and neurodegeneration.

biochemistry↗

bioRxiv2025

Protein Secondary Structure Patterns In Short-Range Cross-Link Atlas

Cross-linking mass spectrometry (XL-MS) has become a powerful tool in structural biology for investigating protein structure, dynamics, and interactomics. However, short-range cross-links, defined as those connecting residues fewer than 20 positions apart, have traditionally been considered less informative and largely overlooked, leaving significant data unexplored in a systematic manner. Here, we present a system-wide analysis of short-range cross-links, demonstrating their intrinsic correlation with protein secondary structure. We introduce the X-SPAN (Cross-link Structural Pattern Analyzer) software, which integrates publicly available XL-MS datasets from system-wide experiments with AlphaFold-predicted protein structures. Our analysis reveals distinct cross-linking patterns that reflect the spatial constraints imposed by secondary structural elements. Specifically, -helices exhibit periodic cross-linking patterns consistent with their characteristic helical pitch, whereas coils and {beta}-strands display nearly monotonic distributions. A context-dependent protein grammar reinforces short-range cross-link specificity. Short-range cross-links can enhance the statistical inference of secondary structures within integrative modeling workflows. Additionally, our work establishes a framework for benchmarking AlphaFolds local prediction accuracy and provides novel quality control criteria for XL-MS experiments. We anticipate that X-SPAN and our short-range cross-link database will serve as a valuable resource for exploring local secondary structure rearrangements and their potential roles in protein function and allosteric regulation.

molecular biology↗