Metal ions restore the proteolytic resistance of denatured conglutin gamma, a lupin seed glycoprotein, by promoting its refolding.
The susceptibility to trypsin of conglutin gamma, a lupin seed glycoprotein affected by this enzyme only when in a non-native conformation, was used to study the effect of Zn(2+) and other metal ions on the structural dynamics of the protein. When acid-treated trypsin-susceptible conglutin gamma was incubated at neutral pH in the presence of Zn(2+), it became resistant to tryptic attack, contrary to the protein treated in the absence of Zn(2+). The time course of this refolding event has been quantitatively evaluated by SDS-PAGE. Amino acid sequencing of the major polypeptide fragments, produced by trypsin before completion of the refolding process, indicated that only a few cleavable bonds were accessible to the enzyme. This suggested that the presence of metal ions affected the pathway of degradation of the protein, by inducing its folding. Among the other metal ions tested, Ni(2+) also promoted the adoption of a trypsin-resistant conformation of conglutin gamma, whereas Mn(2+) and Ca(2+) had only much lower effects. The relevance of these findings for a deeper understanding of the in vivo degradation of plant food proteins and how it is affected by metal ions are discussed.
Duranti
Marcello
M
Dipartimento di Scienze Molecolari Agroalimentari and Istituto di Fisiologia Veterinaria e Biochimica, Università degli Studi di Milano, Milano, Italy. marcello.duranti@unimi.it
Di Cataldo
Annalisa
A
Sessa
Fabio
F
Scarafoni
Alessio
A
Ceciliani
Fabrizio
F
D004351:Drug Resistance; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000494:pharmacology*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008670:Metals / Q000494:pharmacology*; D009532:Nickel / Q000494:pharmacology; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D010940:Plant Proteins / Q000737:chemistry* / Q000378:metabolism; D011489:Protein Denaturation; D017510:Protein Folding; D020539:Sequence Analysis, Protein; D014357:Trypsin / Q000494:pharmacology*
D008670:Metals; D010446:Peptide Fragments; D010940:Plant Proteins; C079483:conglutin-gamma protein, Lupinus angustifolius; D009532:Nickel; D010450:Endopeptidases; D014357:Trypsin