Optimal conditions for the maximum production of alkaline protease by Bacillus subtilis SHmIIIa were evaluated by Plackett-Burman design. Nine process parameters namely, pH, temperature, agitation, inoculum, glucose, peptone, KH 2 PO 4, FeSO 4 and tween 20 at two levels were selected for the design. Out of nine selected parameters, six parameters (pH 9.75, agitation 225 rpm, inoculum 3.5%, glucose 5.5 g/L, peptone 3.5 g/L and KH 2 PO 4 1.5%, have shown significant influence on alkaline protease production. Optimization has resulted in spectacular enhancement in alkaline protease yield from 966 to 36000 EU/mg/ml with 37.26 fold increase.