Time-dependent proteolytic degradation of chicken egg-white proteins by Proteinase K: qualitative and semi-quantitative evidence from 12.5% SDS-PAGE

Egg white is a protein-rich matrix dominated by ovalbumin, ovotransferrin, ovomucoid and lysozyme, several of which are clinically important food allergens. Controlled proteolysis can reshape the molecular-weight (MW) distribution of egg-white proteins and is widely used to generate functional hydrolysates and to attenuate allergenicity. This study qualitatively and semi-quantitatively evaluated the time-dependent digestion of diluted chicken egg white by Proteinase K (PK) using 12.5% sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). Diluted egg white (1 mg/mL) in Tris-HCl-MgCl2 buffer was incubated at 50 °C for 30, 60 and 90 min, without PK or with PK at 200 µg/mL. Reactions were stopped by heating (95 °C, 5 min), and supernatants were resolved on 12.5% SDS-PAGE alongside a 25-245 kDa ladder. A semi-logarithmic ladder calibration (R2 = 0.99 across the 25-100 kDa resolving range) was used to estimate apparent band masses. No-PK controls retained prominent bands at apparent ~95-120 kDa, ~70-78 kDa (ovotransferrin region) and a dominant ~45-50 kDa band (ovalbumin region). PK-treated lanes showed time-dependent loss of these intact bands, with substantial depletion by 60 min and near-complete loss of major Coomassie-detectable proteins by 90 min, whereas heating at 50 °C without PK did not reproduce this depletion. The results demonstrate efficient, enzyme-dependent proteolysis of major egg-white proteins, including the principal allergens ovalbumin and ovotransferrin, supporting PK digestion as a simple model for producing egg-white hydrolysates and a candidate strategy for allergen-epitope reduction that warrants confirmation by densitometry, peptide profiling and IgE-binding assays.