Keratin is insoluble structural protein that is extensively cross-linked by cysteine disulfide bonds, hydrogen bonding, and hydrophobic interatctions. Therefore, keratin is insoluble and not degradable by general protease such as trypsin, pepsin, and papain. Feathers are almost pure keratin protein. Since chicken meat consumption is increasing annually, feather keratin wastes will be increasing. Therefore, they become feather meal used as animal feed after undergoing physical and chemical treatment. But these processes require significant energy and also destroy certain amino acids such as arginine, cystine. The chemical process also causes environmental pollution. Consequently, biodegradation by microorganisms with keratinolytic activity represents an alternative method to improve the nutritional value of feather waste and to prevent environmental pollution. The potential applications of keratinolytic protease include dehairing of leather, cosmetic industry, detergent industry, drug delivery, and prion protein degradation. The aim of this study was to isolate and characterize feather-degrading bacteria with keratinolytic activity.
A strain H10 was isolated from soil at poultry farm and identified as Pseudomonas hibisciola H10. The optimal condition for keratinolytic protease production was investigated using one variable at a time (OVT) method. The optimal culture conditions for keratinolytic activity by the strain H10 were initial pH 10, 25℃ and 200 rpm, respectively. The optimal medium compositions for keratinolytic protease production were glucose 0.15%, beef extract 0.08%, KH2PO4 0.12%, K2HPO4 0.02%,NaCl 0.07%, MgSO47H2O 0.02%, MgCl26H2O 0.03% and 0.2% feather, respectively. The maximum activity of keratinolytic protease was 29.6 U/ml in an optimal condition. Response surface methodology (RSM) was applied to optimize the culture condition; optimal conditions were glucose 0.20%, KH2PO4 0.05% and pH 9, respectively. Under this condition, the strain H10 produced 33.3 U/ml of keratinolytic protease. Characteristics of partially purified enzyme were investigated. The optimum pH and temperature for the enzyme activity were 9 and 30℃ respectively. The enzyme activity was significantly inhibited by PMSF and EDTA, indicating that the enzyme is serine-type metalloprotease. The kerainolytic protease from the strain H10 could degrade various proteinous substrates such as nail, wool, hair, elastin and collagen. The enzyme showed promising result in the dehairing of cow skin. The enzyme could also effectively remove blood strain from the cotton fabric, thus making it suitable to use as an detergent additive. The enzyme degraded nematodes and improved wool quality without using any chemicals. These environment-friendly properties of the keratinolytic protease from the strain H10 showed that the enzyme could be used for various industrial applications.