Production Of Proteases By Alkalophilic Bacteria From A Hot Sprinq At Wondo Genet

Two organisms with high proteolytic activity on casein,rngelatin or haemoglobin were selected out of the bacterial isolatesrnobtained from a hot spring at Wondo Genet. The testrnorganisms were Gram posi ti ve, rod-shaped, spore-forming withrnchains of two or more cells, motile bacteria. Based on thernbiochemical and morphological tests, it has been confirmed thatrnthey belong to the genus Bacillus. Growth of the two isolates wasrnobserved in the pH and temperature ranges of between 5-10, andrn30-55 °C , respectively. Their optimum pH and temperature forrngrowth were 10 and 55°C. Glucose 0.5% and peptone at 0.5%, 0.4%,rnrespectively resulted better yield of biomass and proteasernproduction. 0.5mM Ca +2 , O.lmM Mg+2 , and 1mM P044 increased biomassrnand enzyme production of both isolates. High enzyme productionrnwas observed in the presence of 1mM Cu+2 and 5mM Mn+2 for Bacillusrnsp-A, and O.lmM Zn+2 and 5mM Mn+2 for Bacillus sp-B. The highestrnenzyme activity of both isolates was observed when assayed atrn55°C and pH 9.5. The protease of Baccilus sp-A showed higherrnhydrolytic activity on casein than on gelatin or haemoglobin.rnThe protease of Bacillus sp-B have been found to hydrolyse onlyrngelatin. However, both isolates failed to hydrolyse eggrnalbumin. The two organisms were more efficient in degrading skinrnwhen supplied in the growth medium than feather, horn or hair.rnT high degrading activity on feather than Bacillus sp-B. On thernother hand Bacillus sp-B showed high degrading activity on hornrnthan Bacillus sp-A. The protease of Bacillus sp-A was thermallyrnstable between 60-70 o C, with a half life of 50 min at 60°C andrn65°C, and 40 min at 70°C. The protease of Bacillus sp-B wasrnthermally stable at 60 and 65°C, and the half life was 50 min atrnthese temperatures. However, the enzyme activity of both isolatesrnceased after 60 min of incubation at the temperatures examined.rnviihe least weight lose was recorded on hair. Bacillus sp-A showedrnhigh degrading activity on feather than Bacillus sp-B. On thernother hand Bacillus sp-B showed high degrading activity on hornrnthan Bacillus sp-A. The protease of Bacillus sp-A was thermallyrnstable between 60-70 o C, with a half life of 50 min at 60°C andrn65°C, and 40 min at 70°C. The protease of Bacillus sp-B wasrnthermally stable at 60 and 65°C, and the half life was 50 min atrnthese temperatures. However, the enzyme activity of both isolatesrnceased after 60 min of incubation at the temperatures examined.

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