title: Comparative structure-function characterization of the saposin-like domains from potato, barley, cardoon and Arabidopsis aspartic proteases.
abstract: The present study characterized the aspartic protease saposin-like domains of four plant species, Solanum tuberosum (potato), Hordeum vulgare L. (barley), Cynara cardunculus L. (cardoon; artichoke thistle) and Arabidopsis thaliana, in terms of bilayer disruption and fusion, and structure pH-dependence. Comparison of the recombinant saposin-like domains revealed that each induced leakage of bilayer vesicles composed of a simple phospholipid mixture with relative rates Arabidopsis>barley>cardoon>potato. When compared for leakage of bilayer composed of a vacuole-like phospholipid mixture, leakage was approximately five times higher for potato saposin-like domain compared to the others. In terms of fusogenic activity, distinctions between particle size profiles were noted among the four proteins, particularly for potato saposin-like domain. Bilayer fusion assays in reducing conditions resulted in altered fusion profiles except in the case of cardoon saposin-like domain which was virtually unchanged. Secondary structure profiles were similar across all four proteins under different pH conditions, although cardoon saposin-like domain appeared to have higher overall helix structure. Furthermore, increases in Trp emission upon protein-bilayer interactions suggested that protein structure rearrangements equilibrated with half-times ranging from 52 to 120s, with cardoon saposin-like domain significantly slower than the other three species. Overall, the present findings serve as a foundation for future studies seeking to delineate protein structural features and motifs in protein-bilayer interactions based upon variability in plant aspartic protease saposin-like domain structures.
authors.affiliation: Department of Food Science, Ontario Agricultural College, University of Guelph, Guelph, ON N1G 2W1, Canada.
authors.forename: Brian C
authors.initials: BC
authors.lastname: Bryksa
authors.affiliation: Department of Food Science, Ontario Agricultural College, University of Guelph, Guelph, ON N1G 2W1, Canada.
authors.forename: Douglas A
authors.initials: DA
authors.lastname: Grahame
authors.affiliation: Faculty of Land and Food Systems, University of British Columbia, Vancouver, BC V6T 1Z4, Canada. Electronic address: rickey.yada@ubc.ca.
authors.forename: Rickey Y
authors.initials: RY
authors.lastname: Yada
mesh_terms: D017360:Arabidopsis / Q000201:enzymology*; D057055:Aspartic Acid Proteases / Q000737:chemistry* / Q000502:physiology; D031942:Cynara / Q000201:enzymology*; D001467:Hordeum / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D008051:Lipid Bilayers / Q000737:chemistry; D000072417:Protein Domains; D017433:Protein Structure, Secondary; D049231:Saposins / Q000737:chemistry* / Q000502:physiology; D011198:Solanum tuberosum / Q000201:enzymology*
chemical_list: D008051:Lipid Bilayers; D049231:Saposins; D057055:Aspartic Acid Proteases
keywords: Aspartic protease; Fusion; Membrane disruption; Phospholipid bilayer; Plant-specific insert; Saposin