Immunochemical and structural analysis of pepsin-digested egg white ovomucoid.
Ovomucoid, an egg protein comprising approximately 10% egg white, was digested using the enzyme pepsin, and fragments were isolated by anion-exchange and reverse phase HPLC. Four distinct fragments were identified by analysis with SDS-PAGE, including three large fragments with molecular weights of around 24, 18, and 14 kDa. N- and C-terminal and amino acid sequencing analyses identified the fragments as V134-C186 (domain 3), V21-A133, and A1-A133 (domain 1+2). Further separation and sequencing of the fraction composed of small peptides, to determine their exact makeup and location in the protein, remained to be carried out and identified a peptide G51-Y73. All four fragments showed IgE-binding activity, as measured by ELISA, using human sera from egg-allergic individuals. Little change in the digestibility of ovomucoid by trypsin and chymotrypsin was observed following digestion with pepsin, indicating that pepsin-digested ovomucoid retains its trypsin (protease) inhibitor activities. Reduced carboxymethylated ovomucoid was prepared, and digestion with pepsin produced significantly more peptides than did the digestion of the native ovomucoid, indicating that the disulfide bonds play a significant role in the digestive resistance of ovomucoid. The reduction of ovomucoid enhanced its digestibility and lower allergenicity of the protein.
Kovacs-Nolan
J
J
Department of Food Science, University of Guelph, Guelph, Ontario, Canada N1G 2W1.
Zhang
J W
JW
Hayakawa
S
S
Mine
Y
Y
D004063:Digestion; D004529:Egg White; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D007120:Immunochemistry; D008969:Molecular Sequence Data; D015394:Molecular Structure; D010059:Ovomucin / Q000737:chemistry* / Q000276:immunology*; D010434:Pepsin A / Q000378:metabolism; D010446:Peptide Fragments / Q000032:analysis* / Q000737:chemistry* / Q000276:immunology; D020133:Reverse Transcriptase Polymerase Chain Reaction
D010446:Peptide Fragments; D010059:Ovomucin; D010434:Pepsin A