Biochemical characterization of a surfactant-stable keratinase purified from Proteus vulgaris EMB-14 grown on low-cost feather meal

Abstract

Objectives: The bioaccumulation of keratinous wastes from poultry and dairy industries poses a danger of instability to the biosphere due to resistance to common proteolysis and as such, microbial- and enzyme-mediated biodegradation are discussed.

Results: In submerged fermentation medium, Proteus vulgaris EMB-14 utilized and efficiently degraded feather, fur and scales by secreting exogenous keratinase. The keratinase was purified 14-fold as a monomeric 49 kDa by DEAE-Sephadex A-50 anion exchange and Sephadex G-100 size-exclusion chromatography. It exhibited optimum activity at pH 9.0 and 60 °C and was alkaline thermostable (pH 7.0-11.0), retaining 87% of initial activity after 1 h pre-incubation at 60 °C. The K_m and V_max of the keratinase with keratin azure were respectively 0.283 mg/mL and 0.241 U/mL/min. Activity of P. vulgaris keratinase was stimulated by Ca²⁺, Mg²⁺, Zn²⁺, Na⁺ and maintained in the presence of some denaturing agents, except β-mercaptoethanol while Cu²⁺ and Pb²⁺ showed competitive and non-competitive inhibition with K_i 6.5 mM and 17.5 mM, respectively.

Conclusion: This purified P. vulgaris keratinase could be surveyed for the biotechnological transformation of bioorganic keratinous wastes into valuable products such as soluble peptides, cosmetics and biodegradable thermoplastics.

Keywords: Feather; Inhibition; Keratinase; Poultry waste; Purification; Valorization.