Phylogenetic variations in a novel family of hyperstable apple snail egg proteins: insights into structural stability and functional trends.

The relationship between protein stability and function evolution has not been explored in proteins from natural sources. Here, we investigate the phylogenetic differences of Perivitellin-1 (PV1) a novel family of hyperstable egg carotenoproteins crucial to the reproductive success of Pomacea snails, as they have evolved clade-specific protective functions. We studied P. patula PV1 (PpaPV1) from Flagellata clade eggs, the most basal of Pomacea and compared it with PV1s orthologs from derived clades. PpaPV1 stands as the most stable, with longer unfolding half-life, resistance to detergent unfolding, and therefore higher kinetic stability than PV1s from derived clades. In fact, PpaPV1 is among the most hyperstable proteins described in nature. In addition, its spectral characteristics providing a pale egg coloration, mild lectin activity and glycan specificity are narrower than derived clades. Our results provide evidence indicating large structural and functional changes along the evolution of the genus. Notably, the lectin binding of PpaPV1 is less pronounced, and its glycan specificity is narrower compared to PV1s in the sister Bridgesii clade. Our findings underscore the phylogenetic disparities in terms of structural and kinetic stability, as well as defensive traits like a potent lectin activity affecting the gut morphology of potential predators within the Bridgesii clade or a conspicuous, likely warning coloration, within the Canaliculata clade. This work provides a comprehensive comparison of the structural attributes, stability profiles, and functional roles of apple snail egg PV1s from multiple species within a phylogenetic context. Furthermore, it proposes an evolutionary hypothesis suggesting a trade-off between structural stability and the functional aspects of apple snail's major egg defense protein.