Optimization and Purification of Keratinase from Bacillus anthracis with Dehairing Application

Abstract

The feather waste is treated by land filling or burning which requires a lot of expenditure and also responsible for air, soil and water contamination. The common methods to degrade feathers consume a huge amount of energy and are not good for environment. The biological degradation of feather waste is an environmental friendly method. In this study, bacterial strain MKR-9 was used and identified as Bacillus anthracis by using 16S rRNA sequencing. The keratinase production was carried in keratin medium by using strain MKR-9. During optimization, the maximum keratinase production was observed with 4% inoculum, 1% (w/v) substrate concentration, 40°C temperature, 7.0 pH, 3 days of incubation and presence of methionine. The keratinase production was suppressed by the presence of all used carbon and nitrogen source. The crude keratinase was stable at 40°C and 7.0 pH after pre-incubation of enzyme for 6 h. The crude keratinase can completely remove the goat skin hair after 24 hr of treatment. On the basis of this finding, crude kearatinase mediated process can be employed in lather industry for the removal of hair as it is environmental friendly method. After ammonium sulphate precipitation (80% saturation) and dialysis, the partially purified keratinase was further purified by Sephadex G-75 column chromatography. The protein content, enzyme activity, specific activity, yield and overall purification were measured as 16.38 mg, 9652 U, 589.25 U/mg, 44.89% and 7.93 respectively. The purity of keratinase was confirmed by SDS- PAGE with a single band of 34 kDa.

Keywords

• rrna sequencing
• keratinase
• dehairing
• purification
• sephadex g-75.