Rejina A.K. and Kunhi A.A.M. 2016. Production and Partial Characterization of Keratinase Enzyme from Pseudomonas sp. S-CSR-0004. Abstract Book. National symposium on ‘Biosciences and Technology –Recent Developments and Future Prospects’, SIAS-Centre for Scientific Research, Safi Institute of Advanced Study, Vazhayoor, Malappuram, Kerala, p. 33.

ABSTRACT

Keratinous wastes such as human hair, feathers, horns, hooves, etc., are abundantly available and replenishable. Feathers alone contain over 90% protein formed in huge amount as poultry waste worldwide which leads to feather protein wastage. Keratin is an insoluble fibrous protein and highly stable protein and is resistant to degradation by physical and chemical components. The solution to this problem was made after the discovery of microorganisms which were capable of degrading this insoluble structural protein into value-added products by the production of enzymes (keratinases). Keratinases also possess tremendous industrial applications in various fields. Hence, an attempt was made to isolate a potential keratinase producing bacterium. A positive keratinase producing bacterium was isolated from a soil sample collected from the vicinity of a poultry outlet at Vazhayoor town. The isolated bacterium was identified as Pseudomonas sp. S-CSR- 0004. The bacterium was cultivated in mineral medium (MM) containing feather meal, horn powder, nail & hair, seperatively as substrates. Horn powder was found to be the best substrate. The enzyme activity obtained for horn powder was 108 U/ml followed by feather meal (104 U/ml), nail (100 U/ml) and hair (92 U/ml), whereas the corresponding growth(OD550) was 0.86, 0.30, 0.34, 0.04 and the protein content of the CFS 1, 2.4, 0.5, 2 respectively. Then specific activity however, was in the order nail>feather meal>hair>horn powder, the values being 200, 104, 46, 45 U/mg protein. Optimization of cultural conditions with effect of pH and temperature was studied. The effect of temperature on enzyme activity in CFS was unusual, as it showed two peaks at 35 & 60°C the activity between these two temperatures being much lower. This could be indicative of a contaminating protease. Could be a case of enzyme cannibalism. This fact was further confirmed by the pattern obtained for effect of temperature with casein and keratin as substrate, as two peak temperatures at 40 & 55°C in the case of casein as substrate and single peak at 45°C. The optimum conditions for maximum enzyme production was seen at pH 7.5. The stability of proteinase checked using casein as substrate. The stability of keratinase checked using keratin solution as substrate. Partial purification of enzyme by ammonium sulphate fractionation. 

 

Keywords: Keratinase, characterization, Pseudomanas sp.