National Academy of Agricultural Sciences (NAAS)
|
PRINT ISSN : 2319-7692
Online ISSN : 2319-7706 Issues : 12 per year Publisher : Excellent Publishers Email : editorijcmas@gmail.com / submit@ijcmas.com Editor-in-chief: Dr.M.Prakash Index Copernicus ICV 2018: 95.39 NAAS RATING 2020: 5.38 |
Microbial keratinases have become biotechnologically important since they target the hydrolysis of highly rigid, strongly cross-linked structural polypeptide “keratin” recalcitrant to the commonly known proteolytic enzymes. Soil samples collected from different poultry shops were enriched for keratinase producers on Whole feather agar containing whole feathers as a sole Carbon and Nitrogen source. Among 11 bacterial isolates, 6 isolates showed protease activity. The best keratinase producing bacterium K10 was selected and identified as Bacillus thuringiensis strain Bt407, based on morphological, cultural, biochemical characteristics and 16S rRNA sequence analysis. The isolate exhibited maximum keratinase production (94.52U/ml) in a optimized feather meal medium containing Feather meal (2%), Yeast extract (1%), Starch (1%), MgSO4 6H2O (0.003%), CaCl2 (0.5mM), KH2PO4 (0.5%), K2HPO4 (0.3%), NaCl (0.5%), pH 7, inoculated with 1% v/v pre-grown cell mass and incubated at 37°C on rotary shaker (120 rpm) for 48 hours. The optimum enzyme activity was observed at 55°C and pH 8. Metal ions like Ca+2, Mg+2, and Ba+2 were seen to enhance enzyme activity whereas Cd+2, Cu+2, Fe+3, Hg+2 and Zn+2 were observed to inhibit enzyme activity. Inhibitors such as SDS helped to retain the activity of the enzyme while 2- mercaptoethanol, DMSO and EDTA were seen to inhibit the enzyme activity. The molecular weight of the keratinase was found to be 33kDa by SDS-PAGE method. Zymography was carried out to show protease activity of the keratinase. Depilatory action of keratinase on goat skin was also demonstrated. The applications of the enzyme as a detergent additive and enzyme hydrolyzed feather meal in bacteriological medium as nitrogen source were also studied. Of the tested keratinous materials used as substrates, the production of enzyme was seen to be more in the presence of human nails than human hairs.
Citations |
 |
 |
 |
 |
