This study was conducted to optimize conditions for crude proteases activity from

Bacillus subtilis

GS-1 by Plackett-Burman (PB) design and response surface methodology. Meanwhile

potential applications of proteases in protein hydrolysis were also determined. Three significant variables (NH

4

Cl

initial pH value and cultural temperature) necessary for the enhancement of proteases activity were selected by using PB design. Furthermore

an empirical model was developed to describe the relationships between these tested variables. Maximum enzymatic activity was attained at 372.814 U/mL

with NH

4

Cl at 3.768 g/L

pH at 5.6 and temperature at 22.4 ℃. Results of hydrolysis experiments indicated that the protease more prefer to hydrolysis proteins from animal source than that from plant source. The protease was prone to hydrolyze chemical bond that composed by amino acids of Glu

Val

Ile

Leu

Phe or Arg

and had great potential for applications in protein hydrolysis.