Electronic Thesis/Dissertation


A recombinant Sp185/333 protein, rSpTransformer, from the Purple Sea Urchin has multitasking binding and shape changing abilities Open Access

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The California purple sea urchin, Strongylocentrotus purpuratus, is a relatively long-lived marine invertebrate with ~50 year lifespan. The longevity of this organism is surprising considering it mainly relies on innate immunity, without antibodies or lymphocytes to continuously fend off a broad range of pathogens (bacteria, fungi, viruses and parasites). We use the purple sea urchin to understand the innate immune system with a specific focus on the proteins encoded by the immune gene family, Sp185/333. The amino acid composition of the Sp185/333 proteins provide no folding prediction and suggest they are intrinsically disordered, making functional predictions for these proteins difficult. The encoded proteins are hypothesized to be anti-microbial with putative immune functions based on the sequence diversity among genes, messages and deduced proteins, and the inducible Sp185/333 gene expression upon immune challenge with bacteria and pathogen-associated molecular patterns (PAMPs). A recombinant Sp185/333 protein, rSp0032, shows specific binding towards marine Gram negative bacteria Vibrio diazotrophicus, and Saccharomyces cerevisciae, but not to Bacillus species. rSp0032 also binds to specific PAMPs and transforms from intrinsic disorder to α helix, and consequently is renamed as rSpTransformer (rSpTrf). Sp185/333 proteins are present on the surface and within vesicles of the sea urchin coelomocytes (immune cells) despite the lack of a transmembrane prediction. Protein-phospholipid binding results indicate that rSpTrf binds to phosphatidic acid (PA), a small phospholipid that is a minor lipid in cell membranes. Synthetic liposomes with PA show membrane instability upon presence of rSpTrf, which induces liposome leakage or lysis, budding, fusion and invagination. Given that rSpTrf has multitasking binding abilities towards a range of binding targets, and transforms from disordered to α helix in the presence of binding targets, many or all of the different variants of Sp185/333 proteins may have similar functions with equivalent shape-changing abilities and hence a new name of SpShapeShifter (SpSh). Therefore, we propose that the SpSh immune protein family may be capable of facilitating effective host protection against a broad array of marine pathogens.

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