Molecular characterization and Cryo-EM structure of the hemocyanin of the invasive freshwater snail Pomacea canaliculata

ITUARTE, SANTIAGO; OTERO, LISANDRO H.; BROLA, TABATA; QIU, J; DREON, MARCOS; CHIUMIENTO, IGNACIO; SUN, J; HERAS, HORACIO

San Luis

Congreso; XLVIII Reunión Anual de la Sociedad Argentina de Biofísica; 2019

Sociedad Argentina de Biofísica

Hemocyanins(HC) are the respiratory proteins found in most mollusks andarthropods.Probably due to their huge size and glycan moietiesmolluscan HC can be used as immuno stimulants. Nevertheless,structural knowledge of these proteins is scarce. In this work westudied the molecular assembly, glycosylation pattern and genestructure of the HC from the gastropod P. canaliculata(PcH).Through a proteomic and genomic approach, we identified 4 genesequences of PcH subunits. Each subunit has the typical architectureof a gastropod HC, which is a string of eight globular functionalunits (Fus) of about 50 kDa each. Correspondingly, genes areorganized in eight structurally related coding regions, with linkerintrons of variable length. Of special relevance for evolutionaryconsiderations are introns interrupting regions that encode adiscrete FU, a feature not found in other mollusks. Although thepositioning of the FUs linker introns is highly conserved amongmollusks, introns within FUs show no relationship neither inlocationnor phase. A preliminary low-resolution model of its structure wasachieved by single-particle cryo-electron microscopy. PcH structureshows a cylindrical rearrangement assembled by di, tri, andtetra-decamers with an internal collar structure. PcH isN-glycosylated with oligo- or high mannose and hybrid-typestructures, and complex-typeN-linked glycans, without sialic acid. Itis also decorated by terminal β-N-GlcNAc residuesand non reducingterminal α-GalNAc. This glycosylation profile, with an abundance ofN-linked glycans, is similar to that of Megathura crenulataHC. Inaddition, polyclonalantibodies against PcH do not cross-react withother gastropod HCs, thus highlighting structural differences amongthem and opening a research avenue on the immuno stimulant activityof PcH and the role that carbohydrates play in it.p { margin-bottom: 0.25cm; line-height: 115%; }