In this study, effects of medium components and inoculum size on the protease production by Bacillus sp. EBTA6 that was isolated from a home-made Tarhana sample were investigated. The cell-free supernatant of bacterium cultured on a shaking incubator for 24 h was used to test protease activity as the response. With a total number of 11 factors, 12 different experiments were run and the highest experimental protease activity was measured as 2280.4 U/mL. Results were analyzed statistically by ANOVA and the most efficient factors were detected as yeast extract, dipotassium phosphate, casein, and peptone with a contribution of 93.78, 2.19, 1.96, 1.31%, respectively. For validation of the selected factors, a further experiment was performed by using of yeast extract (9.98 g/L), dipotassium phosphate (1.27 g/L), casein (8.69 g/L), and peptone (9.88 g/L) obtained from the design equation. The experimental response was found as 2411.4 U/mL which was only 5.5% higher than the predicted response showing that the model was applicable.