Production and Partial Characterization of an Extracellular Thermophile Alkaline Protease from a Selected Strain of Bacillus sp Isolated from Abattoir Soil in the North Region of Cameroon

Abstract

The present study aims at screening and isolates thermostable alkaline protease producing bacteria from soil sample in the northern part of Cameroon, and characterizes their enzyme. Samples of soil were collected from four (04) North Cameroonian towns (Ngaoundere, Garoua, Figuil and Maroua). Six (06) isolates showed important proteolytic activity on skim milk agar. The strain N2.3.5 from Ngaoundere showed the largest hydrolysis halo (24 mm) and was selected for protease production. The partial identification of strain N2.3.5 through morphological and biochemical characterization enabled us to classify it as belonging to the genus Bacillus sp. The growth of the strain N2.3.5 and its enzyme production profile showed that the growth of the bacteria was a diauxic type, and that the protease production was not directly associated with the microbial growth. In a medium composed of soya flour, corn flour, yeast extract and at alkaline pH, following the monitoring of each of these parameters by a complete factorial experimental design, the pH value of 10 was found to be significant for an optimal growth. Meanwhile, the corn flour, the soya flour and the yeast extract did not have a consequent influence on the protease production by the bacteria, under the chosen experimental conditions. The extracellular protease produced by Bacillus sp N235 was partially purified using a precipitation with ammonium sulphate, a dialysis and a gel filtration on sephadex G-75. A specific activity of 1516.7 U/mg and a purification yield of 37.63% were obtained. The partially purified enzyme was able to maintain its activity after been heat-treated at 80°C for 30 min. The maximal activity was exhibited at temperature of 80°C and to a pH 12. The enzyme activity and stability was enhanced in presence of the Ca2+.