Structural and kinetic stability of Pomacea egg carotenoproteins

M. Y. PASQUEVICH; H. HERAS

Congreso; XLVII Reunión Anual de la Sociedad Argentina de Biofísica; 2018

The freshwater snails of Pomacea genus deposit pigmented eggs above the water.Pigmentation is provided by abundant oligomeric carotenoproteins which supplynutrients and photoprotection to the growing embryo. Homologous eggcarotenoproteins have evolved different defensive roles. For instance, in theinvasive canaliculata clade, PcOvo and PmPV1, provide bright reddish colors, awarning coloration advertising the presence of toxins in the eggs; on the contrary,in the non-invasive bridgesii clade, PsSC with a pale color (non-warning signal),have a remarkable lectin activity not found in the canaliculata clade. The aim ofour work is to begin to unveil the structure-function evolution of Pomaceacarotenoproteins. We studied the structural stability and the resistance toproteolysis of PpPV1, a pale P. patula carotenoprotein member of the flagellateclade (the most basal of the genus) and compare it with the other clades ofPomacea. The stability of PpPV1 was studied against pH, temperature andguanidinium chloride (GdnHCl), using spectrophotometry and spectrofluorometry;the resistance to proteolysis was determined by proteinase K and in vitrosimulated gastrointestinal digestions. PpPV1 remained stable over temperaturesup to 85°C and over a wide range of pH (4.0-12.0), with slight alterations at pH2.0. The unfolding equilibrium to GdnHCl showed that while 50% of PmPV1 andPcOVO are unfolded at 2.7 M, basal PpPV1 do so at 5.4 M. The oligomer wasresistant to proteolysis, a feature that, together with the resistance to SDSdenaturation, is shared by kinetically stable proteins. The study revealed that allPomacea carotenoproteins share similar resistance to thermal denaturation,extreme pHs and proteolysis. However, PpPV1 has a remarkable greater resistanceto chemical denaturation than its homologs. The loss of stability along evolutioncould have contributed to the gain of new functions, such as acquiring a strongwarning coloration in the most derived species.