Identification and Characterization of a Pepsin- and Chymotrypsin-Resistant Peptide in the α Subunit of the 11S Globulin Legumin from Common Bean (Phaseolus vulgaris L.)

The 11S globulin legumin typically accounts for approximately
3%
of the total protein in common beans (Phaseolus vulgaris). It was previously reported that a legumin peptide of approximately
20 kDa is resistant to pepsin digestion. Sequence prediction suggested
that the pepsin-resistant peptide is located at the C-terminal end
of the α-subunit, within a glutamic acid-rich domain, overlapping
with a chymotrypsin-resistant peptide. Using purified legumin, the
peptide of approximately 20 kDa was found to be resistant to pepsin
digestion in a pH-dependent manner, and its location was determined
by two-dimensional gel electrophoresis and LC-MS-MS. The location
of the chymotrypsin-resistant peptide was confirmed by immunoblotting
with peptide-specific polyclonal antibodies. The presence of a consensus
site for proline hydroxylation and arabinosylation, the detection
of hydroxyproline residues, purification by lectin affinity chromatography,
and a difference in electrophoretic migration between the chymotrypsin-
and pepsin-resistant peptides suggest the presence of a large O-glycan within these peptides.