Bacillus strains isolated under extreme alkaline conditions (Izmir, Turkey), were screened and identified for high alkaline protease activity. Strains with high protease yields were optimized with respect to inoculum concentration, temperature, agitation speed, initial medium pH and incubation time. Three Bacillus strains coded as I18, L18 and L21 showed high potential, for alkaline protease activity (160-222 U/ml) among 85 isolates. The specific growth rates were estimated from the growth curves as 0.49 h −1 for I18, as 0.6 and 0.7 h −1 for L18 and L21, respectively. The optimum temperatures were determined as 30 • C for strain I18 and 37 • C for the strains L18 and L21. Similarly, the optimum agitation speeds were 100 rpm for I18 and 180 rpm for L18 and L21. For all three strains, the optimum inoculation ratio and incubation time, were determined as 5% (v/v) and 96 h, respectively. The optimum initial media pH was found as pH 10 for strain L18 and L21. Bacillus sp. L21 with the highest specific protease activity (60 U/mg protein) and a broader pH range was chosen for further study. The biomass and product yield for this strain was determined as 0.023 g cell/g glucose and 0.021 U/g glucose, respectively. The crude enzyme of this strain was further characterized and was determined as a bleach stable, serine alkaline protease with an optimum temperature of 60 • C and a pH of 11, with a potential to be a candidate for the applications in the detergent industry.

Bacillus sp. L21 was isolated from the by-products of a leather factory (located in Izmir, Turkey) working under extreme alkaline conditions. Its phenotypic and genotypic identifications were completed, and determined as a potential alkaline protease producer. After screening various elements, carbon and nitrogen sources, soybean meal, maltose50, tween80 and the initial pH conditions were chosen as main factors to be used in the experimental design and response surface methodology (RSM) for the optimization of a low cost enzyme producing media for potential use on an industrial scale. The optimized values obtained by the statistical analysis showed that soybean meal at 3.0 g/l, maltose50 between the ranges of 30 and 40 g/l, tween80 at 0.35 g/l and an initial pH of 8.0 gives maximum protease activity. #