The objectives of this study were to establish the proteolytic effects of Staphylococcus aureus during mastitis on economically important milk proteins. Concentrations of milk proteins were determined by capillary electrophoresis in an experimental model using a field strain of S. aureus. The pathogen was inoculated into bacteria-free control milk to imitate proteolysis caused by the pathogen in the mammary gland between milkings. Milk content of caseins (CN) α(S1), α(S2), κ, β(A1), and β(A2) and whey proteins α-lactalbumin and β-lactoglobulin were analyzed initially and after 6 h of incubation. After 6 h, the overall CN content was significantly reduced (21%) in milk inoculated with S. aureus compared with the bacteria-free control milk. S. aureus significantly lowered concentration of α(S1)-CN (2.5%), β(A1)-CN (3%), and β(A2)-CN (5%). S. aureus also hydrolyzed κ-CN into para-κ-CN, with significant reduction of κ-CN (7.4%) as a consequence.