The study reports production of proteolytic enzyme using cattle dung based biodigested slurry as a substrate and fungal inoculants namely Trichoderma harzianum MTCC 792 and Aspergillus sp. (BL4). Fresh cattle dung and biodigested slurry were analysed for pH, proximate (total solids, volatile solids and ash), chemical (cellulose, hemicellulose, lignin and silica) and biochemical (total proteins, protease and nitrogen) composition. Maximum protein content (1.19 mg/ml) was obtained in autoclaved cattle. Maximum protease activity of 73.60U/g was obtained in T. harzianum MTCC 792 inoculated autoclaved biodigested slurry on 4th day while in Aspergillus sp. (BL4) inoculated autoclaved biodigested slurry, maximum activity of 67.33U/g was obtained on 2nd day. Use of biodigested slurry for proteolytic enzyme production is significant with reference to not only disposal of biodigested slurry but also its value addition for industrial applications. The study indicates that cattle dung and biodigested slurry are good substrates for proteolytic enzyme production.