Abstract

A Gram-positive, rod-shaped, spore-forming haloalkaliphilic bacterium designated as NA7 was isolated from the surface of a Helianthemum nummularium root sample obtained from Wadi Natrun in Egypt. Sequence analysis of the 16S rRNA gene revealed a Bacillus haloalkaliphilius strain as the closest match with 99% identity. In a shake flask culture containing 10% NaCl, adjusted to pH 10 and incubated at 37°C, the isolated strain produced thermostable extracellular alkaline protease with relatively stable maximum activity records (0.610-0.625 TU) within a relatively long stationary phase that exceeded 60 h. A 2-level fractional factorial design (Plackett-Burman) was then applied to screen for nutritional and cultivation factors regulating protease production by the isolate and to appraise their effects. Calculated statistical parameters revealed that NaCl and MgSO₄ are the most significant independent variables affecting alkaline protease production by NA7 and suggested a near-optimum culture condition. Verification of this predicted condition resulted in an alkaline protease specific activity record of 509 TU/mg protein with a 1.27 fold increase when compared to the basal medium culture.

Key words: Alkaline protease, Bacillus haloalkaliphilus, Wadi Natrun, haloalkaliphiles.