Abstract
A novel feather-degrading microorganism was isolated from poultry waste, producing a high keratinolytic activity when cultured on broth containing native feather. Complete feather degradation was achieved during cultivation. The bacterium presents potential use for biotechnological processes involving keratin hydrolysis. Chryseobacterium sp. strain kr6 was identified based on morphological and biochemical tests and 16S rRNA sequencing. The bacterium presented optimum growth at pH 8.0 and 30 °C; under these conditions, maximum feather-degrading activity was also achieved. Maximum keratinase production was reached at 25 °C, while concentration of soluble protein was similar at both 25 and 30 °C. Reduction of disulfide bridges was also observed, increasing with cultivation time. The keratinase of strain kr6 was active on azokeratin and azocasein as substrates, and presented optimum pH and temperature of 7.5 and 55 °C, respectively. The keratinase activity was inhibited by 1,10-phenanthroline, EDTA, Hg2+, and Cu2+ and stimulated by Ca2+.
Similar content being viewed by others
References
Atalo K, Gashe BA (1993) Protease production by a thermophilic Bacillus species (P-001A) which degrades various kinds of fibrous proteins. Biotechnol Lett 15:1151–1156
Auld DS (1995) Removal and replacement of metal ions in metallopeptidases. Methods Enzymol 248:228–242
Böckle B, Müller R (1997) Reduction of disulfide bonds by Streptomyces pactum during growth on chicken feathers. Appl Environ Microbiol 63:790–792
Böckle B, Galunski B, Müller R (1995) Characterization of a keratinolytic serine protease from Streptomyces pactum DSM40530. Appl Environ Microbiol 61:3705–3710
Bradbury JH (1973) The structure and chemistry of keratin fibers. Adv Prot Chem 67:111–211
Elmayergi HH, Smith RE (1971) Influence of growth of Streptomyces fradiae on pepsin-HCl digestibility and methionine content of feather meal. Can J Microbiol 17:1067–1072
Felsenstein J (1989) PHYLIP – Phylogeny Inference Package (version 3.2). Cladistics 5:164–166
Grzywnowicz G, Lobarzewski J, Wawrzkiewicz K, Wolski T (1989) Comparative characterization of proteolytic enzymes from Trichophyton gallinae and Trichophyton verrucosum. J Med Vet Mycol 27:319–328
Jooste PJ, Britz TJ (1986) The significance of Flavobacteria as proteolytic psychotrophs in milk. Milchwissenschaft 41:618–621
Kaul S, Sumbali G (1997) Keratinolysis by poultry farm soil fungi. Mycopathologia 139:137–140
Kim JM, Lim WJ, Suh HJ (2001) Feather-degrading Bacillus species from poultry waste. Process Biochem 37:287–291
Kunert J (1989) Biochemical mechanism of keratin degradation by the actinomycete Streptomyces fradiae and the fungus Microsporum gypseum: a comparison. J Basic Microbiol 29:597-604
Kunert J, Stransky Z (1988) Thiosulfate production from cysteine by the keratinophilic prokatyote Streptomyces fradiae. Arch Microbiol 150:600–601
Kushwaha RKS (1983) The in vitro degradation of peacock feathers by some fungi. Mykosen 26:324–326
Lee GG, Ferket PR, Shih JCH (1991) Improvement of feather digestibility by bacterial keratinase as a feed additive. FASEB J 59:1312.
Letourneau F, Soussotte V, Bressollier P, Branland P, Verneuil B (1998) Keratinolytic activity of Streptomyces sp. SK1–02: a new isolated strain. Lett Appl Microbiol 26:77–80
Lin X, Kelemen DW, Miller ES, Shih JCH (1995) Nucleotide sequence and expression of kerA, the gene encoding a keratinolytic protease of Bacillus licheniformis PWD-1. Appl Environ Microbiol 61:1469–1474
Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurement with the Folin phenol reagent. J Biol Chem 193:267–275
Lucas FS, Broennimann O, Febbrarro I (2003) High diversity among feather-degrading bacteria from a dry meadow soil. FEMS Microbiol Ecol (in press)
MacFaddin JF (2000) Biochemical tests for identification of medical bacteria. Lippincott, Williams and Wilkins, Baltimore
Nam GW, Lee DW, Lee HS, Lee NJ, Kim BC, Choe EA, Hwang JK, Suhartono MT, Pyun YR (2002) Native-feather degradation by Fervidobacterium islandicum AW-1, a newly isolated keratinase-producing thermophilic anaerobe. Arch Microbiol 178:538–547
Noval JJ, Nickerson WJ (1959) Decomposition of native keratin by Streptomyces fradiae. J Bacteriol 77:251–263
Onifade AA, Al-Sane NA, Al-Musallam AA, Al-Zarban S (1998) Potentials for biotechnological applications of keratin-degrading microorganisms and their enzymes for nutritional improvement of feathers and other keratins as livestock feed resources. Biores Technol 66:1–11
Osborn AM, Moore ERB, Timmis KN (1999) An evaluation of terminal-restriction fragment length polymorphisme (T-RFLP) analysis for the study of microbial community structure and dynamics. Environ Microbiol 2:39–45
Palleroni NJ (1984) Facultatively anaerobic Gram-negative aerobic rods and cocci. In: Krigg NR (ed) Bergey's Manual of Systematic Bacteriology, vol 1. Williams & Wilkins, Baltimore, pp 140–407
Papadopoulos MC, El Boushy AR, Roodbeen AE, Ketelaars EH (1986) Effects of processing time and moisture content on amino acid composition and nitrogen characteristics of feather meal. Anim Feed Sci Technol 14:279–290
Parry DAD, North ACT (1998) Hard α-keratin intermediate filament chains: substructure of the N-and C-terminal domains and the predicted structure and function of the C-terminal domains of type I and type II chains. J Struct Biol 122:67–75
Reichenback H (1992) The Order Cytophagales. In: Balows A, Trüper HD, Dworkin M, Harder W, Schleifer KH (eds) The prokayiotes – a handbook of the biology of bacteria: ecophysiology, isolation, identification, applications. Springer, Berlin Heidleberg New York, pp 3631–3675
Sangali S, Brandelli A (2000) Feather keratin hydrolysis by a Vibrio sp. strain kr2. J Appl Microbiol 89:735–743
Santos RMD, Firmino AAP, Sá CM, Felix CR(1996) Keratinolytic activity of Aspergillus fumigatus Fresenius. Curr Microbiol 33:364–370
Shih JCH (1993) Recent development in poultry waste digestion and feather utilization—a review. Poultry Sci 72:1617–1620
Sorimachi H, Ishiura S, Suzuki K (1997) Structure and physiological function of calpains. Biochem J 328:721–732
Suh HJ, Lee HK (2001) Characterization of a keratinolytic serine protease from Bacillus subtilis KS-1. J Prot Chem 20:165–169
Sullivan MX, Hess WC, Howard HW (1942) The quantitative estimation of both cystine and cysteine in mixture. J Biol Chem 145:621–624
Tarentino AL, Quinones G, Grimwood BG, Hauer CR, Plummer TH (1995) Molecular cloning and sequence analysis of flavastascin: an O-glycosylated prokaryotic zinc metalloendopeptidase. Arch Biochem Biophys 319:281–285
Tomarelli RM, Charney J, Harding ML (1949) The use of azoalbumin as a substrate in the colorimetric determination of peptic and tryptic activity. J Lab Clin Med 34:428–433
Vandamme P, Bernardet, J.F., Segers P, Kerters K, Holmes B (1994) New perspective in the identification of flavobacteria: description of Chryseobacterium gen nov, Bergeyella gen nov, and Empedobacter nom ver. Int J Syst Bacteriol 44:827–831
Venter H, Osthoff G, Litthauer D (1999) Purification and characterization of a metalloprotease from Chryseobacterium indologenes Ix9a and determination of the amino acid specificity with electrospray mass spectrometry. Prot Expres Purif 15:282–295
Wang X, Parsons CM (1997) Effect of processing systems on protein quality of feather meal and hog hair meals. Poultry Sci 76:491–496
Williams CM, Richter CS, MacKenzie JM, Shih JCH (1990) Isolation, identification, and characterization of a feather-degrading bacterium. Appl Environ Microbiol 56:1509–1515
Williams CM, Lee CG, Garlich JD, Shih JCH (1991) Evaluation of a bacterial feather fermentation product, feather-lysate, as a feed protein. Poultry Sci 70:85–94
Yamaguchi S, Yokoe M (2000) A novel protein-deamidating enzyme from Chryseobacterium proteolyticum sp. nov, a newly isolated bacterium from soil. Appl Environ Microbiol 66:3337–3343
Yamauchi K, Yamauchi A, Kusunoki T, Khoda A, Konishi Y (1996) Preparation of stable aqueous solutions of keratins, and physicochemical and biodegradational properties of films. J Biomed Mat Res 31:439–444
Zaghloul TI (1998) Cloned Bacillus subtilis alkaline protease (aprA) gene showing high level of keratinolytic activity. Appl Biochem Biotechnol 70/72:199–205.
Acknowledgements
A.R. is the recipient of a M.Sc. fellowship from CNPq. A.B. is Research Fellow of CNPq, Brazil. F.L. P.H. acknowledges the Fondation de Famille Sandoz for financing his research (subsidy to P. Heeb). We thank Jitka Lipka for running our samples at the sequencing facility of the Institute of Ecology (University of Lausanne).
Author information
Authors and Affiliations
Corresponding author
Rights and permissions
About this article
Cite this article
Riffel, A., Lucas, F., Heeb, P. et al. Characterization of a new keratinolytic bacterium that completely degrades native feather keratin. Arch Microbiol 179, 258–265 (2003). https://doi.org/10.1007/s00203-003-0525-8
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s00203-003-0525-8