Lupin storage proteins: targeted proteolysis and unforeseen functionalities

State of art and aim: The frequent presence of twin and multiple arginine residues in the amino acid sequence of several storage proteins and the occurrence of a -R-R- endopeptidase in Lupinus albus, L. seed (Magni et al) support the hypothesis that these residues are cleavage-prone sites for selective degradation. Whether such specific cleavages take place in the plant kingdom leading to the liberation of biologically active peptides, as they do in mammalian cells (Becker KL), is not known yet. The 11S storage globulin binds to a metal affinity chromatography matrix and the role of two unusual stretches of contiguous histidine residues at the C-terminusof the 11S globulin acidic chains, was hypothesized. Being this region rich in R-R motifs too, the protein was incubated with the lupin seed R-Rendopeptidase to confirm the metal binding activity of this his-rich peptide (Capraro et al). Results and discussion: Upon incubation with the enzyme, the loss of metal binding capacity paralleled that of the anti-his-tag reactive polypeptides. The recovered small proteolytic fragment was analyzed by mass spectrometry and N-terminal sequencing and found to correspond to the 24-mer region cleaved off at twin arginine residues and containing the natural his-tag-like region. Similarly, when lupin seeds were germinated for a few days, the his-tag containing 11S globulin chain was converted into a form devoid of such region, suggesting that this mechanism is part of the natural and complex breakdown of the storage proteins during germination. The liberation of a metal binding peptide at early stages of germination can play a number of physiological roles in the plantlet. This work supports the hypothesis of an orderly proteolytic deconstruction of storage proteins during germination rather than an extensive, disordered breakdown and also add preliminary insight on the potential physiological role of the transiently liberated peptide fragments. Moreover, the present findings open new perspectives in the identification of cryptic bioactivities related to seed ontogenesis and protection originating prior to complete breakdown with a a mammalian-like specific proteolytic event..