title: Susceptibility of lupin gamma-conglutin, the plasma glucose-lowering protein of lupin seeds, to proteolytic enzymes.
abstract: Lupin seed gamma-conglutin, orally administered to animal models, has been shown to display glucose-controlling properties. Therefore, we have addressed the study of gamma-conglutin susceptibility to proteolytic enzymes in vitro as the basis to unveil its metabolic fate in the body. Pepsin treatment at pH 2.0 and 3.0 caused extensive proteolytic breakdown, while at pH 4.0, where pepsin is minimally active, gamma-conglutin was unaffected. Aliquots of the pepsin-treated protein were further incubated with pancreatin at neutral pH. If the protein backbone was already cleaved by pepsin action, then the breakdown by pancreatin was almost complete; alternatively, pancreatin did not affect at all gamma-conglutin polypeptide chain. This was not due to an inhibitory activity of gamma-conglutin, because co-incubation with casein showed complete breakdown of the milk protein. Furthermore, gamma-conglutin was incubated with bromelain, a proteinase effective between pH 4.0 and 7.0. A sharp transition from the uncleavable to the fully cleavable form of gamma-conglutin was observed below pH 4.25. Therefore, it was concluded that (i) gamma-conglutin is resistant to proteolysis at pH greater than 4.0, likely because of a compact native conformation, (ii) an acidic pH renders the protein susceptible to proteases, suggesting the occurrence of a trans conformation, which has also been observed by circular dichroism spectral analysis, and (iii) the protein undergoes an "all or none" degradation pathway, regardless of the enzyme used.
authors.affiliation: Department of AgriFood Molecular Sciences, Universita degli Studi di Milano, Via Celoria, 2 I-20133 Milan, Italy.
authors.forename: Jessica
authors.initials: J
authors.lastname: Capraro
authors.forename: Chiara
authors.initials: C
authors.lastname: Magni
authors.forename: Alessio
authors.initials: A
authors.lastname: Scarafoni
authors.forename: Marcello
authors.initials: M
authors.lastname: Duranti
mesh_terms: D001963:Bromelains / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D007004:Hypoglycemic Agents / Q000378:metabolism*; D031297:Lupinus / Q000737:chemistry*; D010194:Pancreatin / Q000378:metabolism; D010434:Pepsin A / Q000378:metabolism; D010447:Peptide Hydrolases / Q000378:metabolism*; D010940:Plant Proteins / Q000378:metabolism*; D055314:Seed Storage Proteins / Q000378:metabolism*; D012639:Seeds / Q000737:chemistry*
chemical_list: D007004:Hypoglycemic Agents; D010940:Plant Proteins; D055314:Seed Storage Proteins; D010194:Pancreatin; D001963:Bromelains; D010447:Peptide Hydrolases; D010434:Pepsin A