Soloka Mabika Armel Faly1, Rachel Moyen1,2,  Etienne Nguimbi*1,2, Gabriel Ahombo, Raoul Ampa1,2,  Aimé Christian Kayath1,3, Alain Vouidibio1,2, Cyr Jonas Morabandzaand Simon Charles Kobawila1

1Cellular and Molecular Biology Laboratory, Faculty of Sciences and Techniques, Marien Ngouabi University, Brazzaville, Republic of Congo.
2Unit of Microbiology, Molecular Biology and Bioinformatics, Faculty of Sciences and Techniques, Marien Ngouabi University, Brazzaville, Republic of Congo.
3Institute of Research in Exact and Natural Sciences , Ministry of Research and Technologic
innovation, Brazzaville, Republic of Congo.

Abstract

Two Bacillus strains isolated from Ntobambodi  :Bacillus megaterium (B.me NM 02)  Bacillus licheniformis(B.li NM01),has shown a significant proteolytic activity in the caseinolytic character actively degrading casein.In the present work we set optimization of growth and enzyme production conditions. We therefore cultured strains in LB medium, with time computed the OD value for growth. Using containing-skimmed milk petri dishes, we measure the diameter of the formed halo with sample. Several parameters have been optimized: temperature, pH, various types of media, carbon and nitrogen sources. In both strains, growth is possible from 25 to 60 ° C with an optimum temperature at 37 ° C for B.li and at 35°C for B.me. Enzyme production was observed from 25 to 55 ° C with an optimum temperature of 30 ° C for B.li. Enzyme production was observed from 25 to 50 ° C with an optimum temperature of 35 ° C in B.me. For pH, growth and enzyme production can be at 5, 7 and 9 with an optimum at 7 for both strains. The LB medium is better for growth and enzyme production than TSB for B.li and B.me. Among the carbon sources used, fructose is better for growth after 48 hours of incubation in both strains (B.li: 0,93±0,001, B.me: 0,928±0,002), but for enzyme production fructose remains the best carbon source for  B.li (14,33±1,24) , while starch is the best for B.me (14.66±1,24). Concerning nitrogen sources, in both strains the best source of growth is the yeast extracts (B.li: 0,969±0,015, B.me:0,952±0,01). For enzyme production, the two sources can be used forB.li.(14,333±1,247) but for  B .me (14,333±0,471) only the yeast extracts is the  best  as well as for growth and enzyme production. Furthermore, in both strains the produced enzyme was partially purified by using ammonium sulfate precipitation, and SDS-PAGE has been hold, profiles of specific bands are useful to give more information and differentiate the two strains.

Keywords: Optimization, caseinolytic enzyme, Ntobambodi, Bacillus licheniformis, Bacillus megaterium.