Substrate specificity characterization of a thermostable keratinase from Pseudomonas aeruginosa KS-1

Abstract

A feather-degrading strain of Pseudomonas aeruginosa KS-1 was used in the present study. Its crude cell-free fermentation broth completely degraded chicken feather within 12 h, in the absence of disulphide reductase activity. Keratinase from its extracellular broth was purified and characterized, assuming that it would be a potential beta-keratin-degrading enzyme with prospective applications in degradation of beta-plaques of prions. The keratinase was purified by using Q-Sepharose anion exchange chromatography and its molecular weight, as determined by SDS-PAGE analysis, was 45 kDa. It was an alkaline, serine protease with pH and temperature optima of 9 and 60 degrees C, respectively. The enzyme was highly thermostable with a t (1/2) > 2 h at 80 degrees C and had a very high K to C (keratinolytic to caseinolytic) ratio of 2.5. Besides feather keratin, it also hydrolyzed a variety of other complex substrates including fibrin, gelatin and meat protein. Its activity on synthetic substrates revealed that it efficiently cleaves them in the order phenylalanine > lysine > alanine > leucine p-nitroanilides. It also cleaved insulin B chain between Val(12)-Glu(13), Ala(14)-Leu(15), Gly(20)-Glu(21) and Arg(22)-Gly(23) residues.

Metabolism of Chicken Feathers and Concomitant Electricity Generation by Pseudomonas aeruginosa by Employing Microbial Fuel Cell (MFC)

“…It indicates that bacterial and fungal strains exhibit varying keratinolytic potential. The strain SDS3 showed a slow rate of degradation as compared with P. aeruginosa strain KS-1, which showed high rates of feather degradation [27]. These differences can be attributed to metabolic versatility between isolates growing in different environmental conditions.…”

“…In this study, strain SDS3 was employed for metabolism of chicken feathers. Strain SDS3 shows a slow rate of feather metabolism as compared to rapid feather degrading strain KS-1 [27]. This suggests that the use of strain SDS3 in MFC is advantageous because a slow rate of feather degradation will eventually increase the duration of stationary phase of MFC and will lead to greater electricity production over a large time period.…”

Identification and characterisation of a Bacillus licheniformis strain with profound keratinase activity for degradation of melanised feather

“…The latter suggests that the presence of a consortium of enzymes may be required to enhance feather keratin degradation (Sharma and Gupta 2010). In contrast, the keratinase of B.…”

“…fungi (Gradisar et al 2005;Ismail et al 2012), actinomycetes (Bressollier et al 1999), Streptomyces (Tapia and Simoes 2008), Bacillus (Williams et al 1990) and Pseudomonas (Sharma and Gupta 2010) species have been isolated from various environmental sources such as soil; poultry farm wastes and raw feather. The great potential for keratinase in the biotechnology industry demands that the most efficient and environmentally friendly sources are identified and exploited for maximum production.…”

Microbial keratinases: characteristics, biotechnological applications and potential.

“…In prokaryotic cells, sulphitolysis can be achieved by the production of disulphide reductases, release of sulphite and thiosulphate (Kunert 1989;Ramnani et al 2005) or a cell-bound redox system Sharma and Gupta 2010a). Disulphide reductases produced by a number of microbes have been shown to effectively reduce the disulphide bonds, specifically: Streptomyces pactum (Böckle et al 1995), Vibrio Kr 2 (Sangali and Brandelli 2000), StenotrophomonasD-1 and S. maltophilia BBE11-1 sp.…”