Microbial keratinases and their prospective applications: an overview

  • Rani GuptaEmail author
  • Priya Ramnani


Microbial keratinases have become biotechnologically important since they target the hydrolysis of highly rigid, strongly cross-linked structural polypeptide “keratin” recalcitrant to the commonly known proteolytic enzymes trypsin, pepsin and papain. These enzymes are largely produced in the presence of keratinous substrates in the form of hair, feather, wool, nail, horn etc. during their degradation. The complex mechanism of keratinolysis involves cooperative action of sulfitolytic and proteolytic systems. Keratinases are robust enzymes with a wide temperature and pH activity range and are largely serine or metallo proteases. Sequence homologies of keratinases indicate their relatedness to subtilisin family of serine proteases. They stand out among proteases since they attack the keratin residues and hence find application in developing cost-effective feather by-products for feed and fertilizers. Their application can also be extended to detergent and leather industries where they serve as specialty enzymes. Besides, they also find application in wool and silk cleaning; in the leather industry, better dehairing potential of these enzymes has led to the development of greener hair-saving dehairing technology and personal care products. Further, their prospective application in the challenging field of prion degradation would revolutionize the protease world in the near future.


Bovine Spongiform Encephalopathy Nocardiopsis Keratinolytic Activity Feather Meal Keratinase Production 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.



The authors thank the Department of Biotechnology, Government of India, New Delhi, for granting financial assistance through a project on feed enzymes (Sanction no. BT/03/06/2000-PID) and the Council of Scientific and Industrial Research, Government of India, New Delhi, for financial support to P. Ramnani in the form of a Senior Research Fellowship.


  1. Akhtar W, Edwards HGM (1997) Fourier-transform Raman spectroscopy of mammalian and avian keratotic biopolymers. Spectrochim Acta 53:81–90Google Scholar
  2. Allpress JD, Mountain G, Gowland PC (2002) Production, purification and characterization of an extracellular keratinase from Lysobacter NCIMB 9497. Lett Appl Microbiol 34:337–342CrossRefGoogle Scholar
  3. Anbu P, Gopinath SCB, Hilda A, Lakshmipriya T, Annadurai G (2005) Purification of keratinase from poultry farm isolate Scopulariopsis brevicaulis and statistical optimization of enzyme activity. Enzyme Microb Technol 36:639–647CrossRefGoogle Scholar
  4. Apple JK, Boger CB, Brown DC, Maxwell CV, Friesen KG, Roberts WJ, Johnson ZB (2003) Effect of feather meal on live animal performance and carcass quality and composition of growing–finishing swine. J Anim Sci 81:172–181Google Scholar
  5. Bernal C, Vidal L, Valdivieso E, Coello N (2003) Keratinolytic activity of Kocuria rosea. World J Microbiol Biotechnol 19:255–261CrossRefGoogle Scholar
  6. Betzel C, Dauter Z, Dauter M, Ingelman M, Papendorf G, Wilson KS, Branner S (1988) Crystallization and preliminary X-ray diffraction studies of an alkaline protease from Bacillus lentus. J Mol Biol 204:803–804CrossRefGoogle Scholar
  7. Bockle B, Muller R (1997) Reduction of disulfide bonds by Streptomyces pactum during growth on chicken feathers. Appl Environ Microbiol 63:790–792Google Scholar
  8. Bockle B, Galunsky B, Muller R (1995) Characterization of a keratinolytic serine proteinase from Streptomyces pactum DSM 40530. Appl Environ Microbiol 61:3705–3710Google Scholar
  9. Bressollier P, Letourneau F, Urdaci M, Verneuil B (1999) Purification and characterization of a keratinolytic serine proteinase from Streptomyces albidoflavus. Appl Environ Microbiol 65:2570–2576Google Scholar
  10. Brutt EH, Ichida JM (1999) Keratinase produced by Bacillus licheniformis. US Patent 5,877,000Google Scholar
  11. Cheng SW, Hu HM, Shen SW, Takagi H, Asano M, Tsai YC (1995) Production and characterization of a feather degrading Bacillus licheniformis PWD-1. Biosci Biotechnol Biochem 59:2239–2243CrossRefGoogle Scholar
  12. Chitte RR, Nalawade VK, Dey S (1999) Keratinolytic activity from the broth of a feather-degrading thermophilic Streptomyces thermoviolaceus strain SD8. Lett Appl Microbiol 28:131–136CrossRefGoogle Scholar
  13. Choi JM, Nelson PV (1996) Developing a slow-release nitrogen fertilizer from organic sources. II. Using poultry feathers. J Am Hort Sci 121:634–638Google Scholar
  14. De Toni CH, Richter MF, Chagas JR, Henriques JAP, Termignoni C (2002) Purification and characterization of an alkaline serine endopeptidase from a feather-degrading Xanthomonas maltophila strain. Can J Microbiol 48:342–348CrossRefGoogle Scholar
  15. Dozie INS, Okeke CN, Unaeze NC (1994) A thermostable, alkaline-active, keratinolytic proteinase from Chrysosporium keratinophilum. World J Microbiol Biotechnol 10:563–567CrossRefGoogle Scholar
  16. El-Naghy MA, El-Ktatny MS, Fadl-Allah EM, Nazeer WW (1998) Degradation of chicken feathers by Chrysosporium georgiae. Mycopathologia 143:77–84CrossRefGoogle Scholar
  17. Evans KL, Crowder J, Miller ES (2000) Subtilisins of Bacillus spp. hydrolyze keratin and allow growth on feathers. Can J Microbiol 46:1004–1011CrossRefGoogle Scholar
  18. Farag AM, Hassan MA (2004) Purification, characterization and immobilization of a keratinase from Aspergillus oryzae. Enzyme Microb Technol 34:85–93CrossRefGoogle Scholar
  19. Friedrich AB, Antranikian G (1996) Keratin degradation by Fervidobacterium pennavorans, a novel thermophilic anaerobic species of the order thermotogales. Appl Environ Microbiol 62:2875–2882Google Scholar
  20. Friedrich J, Kern S (2003) Hydrolysis of native proteins by keratinolytic protease of Doratomyces microsporus. J Mol Catal 21:35–37CrossRefGoogle Scholar
  21. Friedrich J, Gradisar H, Mandin D, Chaumont JP (1999) Screening fungi for synthesis of keratinolytic enzymes. Lett Appl Microbiol 28:127–130CrossRefGoogle Scholar
  22. Gassessse A, Kaul RH, Gashe BA, Mattiasson B (2003) Novel alkaline proteases from alkalophilic bacteria grown on chicken feather. Enzyme Microb Technol 32:519–524CrossRefGoogle Scholar
  23. Gousterova A, Braikova D, Goshev I, Christov P, Tishinov K, Tonkova VE, Haertle T, Nedkov P (2005) Degradation of keratin and collagen containing wastes by newly isolated thermoactinomycetes or by alkaline hydrolysis. Lett Appl Microbiol 40:335–340CrossRefGoogle Scholar
  24. Gunkel FA and Gassen HG (1989) Proteinase K from Tritirachium album Limber: characterization of the chromosomal gene and expression of the cDNA in E. coli. Eur J Biochem 179:185–194CrossRefGoogle Scholar
  25. Gradisar H, Kern S, Friedrich J (2000) Keratinase of Doratomyces microsporus. Appl Microbiol Biotechnol 53:196–200CrossRefGoogle Scholar
  26. Gregg R (2002). From feathers to degradable plastic. BRI: end of mad cows disease. Triangle Tech Journal (
  27. Gupta R, Beg QK, Lorenz P (2002) Bacterial alkaline proteases: molecular approaches and industrial applications. Appl Microbiol Biotechnol 59:15–32CrossRefGoogle Scholar
  28. Hadas A, Kautsky L (1994) Feather meal, a semi-slow-release nitrogen fertilizer for organic farming. Fertil Res 38:165–170CrossRefGoogle Scholar
  29. Huang Q, Peng Y, Li X (2003) Purification and characterization of an extracellular alkaline serine protease with dehairing function from Bacillus pumilis. Curr Microbiol 43:169–173CrossRefGoogle Scholar
  30. Ignatova Z, Gousterova A, Spassov G, Nedkov P (1999) Isolation and partial characterization of extracellular keratinase from a wool degrading thermophilic actinomycete strain Thermoactinomyces candidus. Can J Microbiol 45:217–222CrossRefGoogle Scholar
  31. Ikemura H, Tagaki H, Inouye M (1987) Requirement of pro-sequence for the production of active subtilisin E in Eschericia coli. J Biol Chem 262:7859–7864Google Scholar
  32. Jacobs I, Eliasson M, Uhlen M, Flock JI (1985) Cloning, sequencing and expression of subtilisin Carlsberg from Bacillus licheniformis. Nucleic Acids Res 13:8913–8926CrossRefGoogle Scholar
  33. Jang JS, Kang DO, Chun MJ, Byun SM (1992) Molecular cloning of Subtilisin J gene from Bacillus stearothermophilus and its expression in Bacillus subtilis. Biochem Biophys Res Commun 184:277–282CrossRefGoogle Scholar
  34. Kaul S, Sambali G (1999) Production of extracellular keratinases by keratinophilic fungal species inhabiting feathers of living poultry birds (Gallus domesticus): a comparison. Mycopathologia 146:19–24CrossRefGoogle Scholar
  35. Kim JM, Lim WJ, Suh HJ (2001) Feather-degrading Bacillus species from poultry waste. Process Biochem 37:287–291CrossRefGoogle Scholar
  36. Kim JS, Kluskens LD, de Vos WM, Huber R, van der Oost J (2004) Crystal structure of fervidolysin from Fervidobacterium pennivorans, a keratinolytic enzyme related to subtilisin. J Mol Biol 335:787–797CrossRefGoogle Scholar
  37. Kluskens LD, Voorhorst WGB, Siezen RJ, Schwerdtfeger RM, Antranikian G, van der Oost J, de Vos WM (2002) Molecular characterization of fervidolysin, a subtilisin-like serine protease from the thermophilic bacterium Fervidobacterium pennivorans. Extremophiles 6:185–194CrossRefGoogle Scholar
  38. Kreplak L, Doucet J, Dumas P, Briki F (2004) New aspects of the α-helix to β-sheet transition in stretched hard α-keratin fibers. Biophys J 87:640–647CrossRefGoogle Scholar
  39. Kurihara M, Markland FS, Smith EL (1972) Subtilisin amylosaccharolyticus. III. Isolation and sequence of the chymotryptic peptides and the complete amino acid sequence. J Biol Chem 247:5617–5631Google Scholar
  40. Langeveld JPM, Wang JJ, Van de Wiel DFM, Shih GC, Garssen GJ, Bossers A, Shih JCH (2003) Enzymatic degradation of prion protein in brain stem from infected cattle and sheep. J Infect Dis 188:1782–1789CrossRefGoogle Scholar
  41. Lee GG, Ferket PR, Shih JCH (1991) Improvement of feather digestibility by bacterial keratinase as a feed additive. FASEB J 59:1312Google Scholar
  42. Leo WD, van Raamsdonk, Vancutsem J, Zegers J, Frick G, Jorgenson JS, Pinckaers V, Bosch J, Severin IP (2004) The microscopic detection of animal proteins in feeds. Biotechnol Agron Soc Environ 8:241–247Google Scholar
  43. 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 26:77–80CrossRefGoogle Scholar
  44. Lin X, Lee CG, Casale ES, Shih JCH (1992) Purification and characterization of a keratinase from a feather-degrading Bacillus licheniformis strain. Appl Environ Microbiol 58:3271–3275Google Scholar
  45. Lin X, Kelemen DW, Miller ES, Shih JCH (1995) Nucleotide sequence and expression of ker A, the gene encoding a keratinolytic protease of Bacillus licheniformis PWD-1. Appl Environ Microbiol 61:1469–1474Google Scholar
  46. Lin X, Shih JCH, Swaisgood HE (1996) Hydrolysis of feather keratin by immobilized keratinase. Appl Environ Microbiol 62:4273–4275Google Scholar
  47. Lin X, Wong SL, Miller ES, Shih JCH (1997) Expression of Bacillus licheniformis PWD-1 keratinase gene in Bacillus subtilis. J Ind Microbiol Biotechnol 19:134–138CrossRefGoogle Scholar
  48. Lin X, Inglis GD, Yanke LJ, Cheng KJ (1999) Selection and characterization of feather degrading bacteria from conola meal compost. J Ind Microbiol Biotechnol 23:149–153CrossRefGoogle Scholar
  49. Lucas FS, Broennimann O, Febbraro I, Heeb P (2003) High diversity among feather-degrading bacteria from a dry meadow soil. Microb Ecol 45:282–290CrossRefGoogle Scholar
  50. Macedo AJ, da Silva WOB, Gava R, Driemeier D, Henriques JAP, Termignoni C (2005) Novel keratinase from Bacillus subtilis S14 exhibiting remarkable dehairing capabilities. Appl Environ Microbiol 71:594–596CrossRefGoogle Scholar
  51. Manczinger L, Rozs M, Vagvolgyi Cs, Kevei F (2003) Isolation and characterization of a new keratinolytic Bacillus licheniformis strain. World J Microbiol Biotechnol 19:35–39CrossRefGoogle Scholar
  52. Mitsuiki S, Ichikawa M, Oka T, Sakai M, Moriyama Y, Sameshima Y, Goto M, Furukawa K (2004) Molecular characterization of a keratinolytic enzyme from an alkaliphilic Nocardiopsis sp. TOA-1. Enzyme Microb Technol 34:482–489CrossRefGoogle Scholar
  53. Mohamedin AH (1999) Isolation, identification and some cultural conditions of a protease producing thermophilic Streptomyces strain grown on chicken feathers as a substrate. Int Biodeterior Biodegrad 43:13–21CrossRefGoogle Scholar
  54. Mukhopadhyay RP, Chandra AL (1990) Keratinase of a streptomycete. Indian J Exp Biol 28:575–577Google Scholar
  55. Musahl C, Aguzzi A (2000) Prions. In: Lederberg J (ed) Encyclopedia of microbiology, 2nd edn. Academic, San Diego, pp 809–823Google Scholar
  56. Nakamura T, Yamagata Y, Ichishima E (1992) Peptide synthesis of proteases in organic solvents: medium effect on substrate specificity. Enzyme Microb Technol 14:842–847CrossRefGoogle Scholar
  57. 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 178:538–547CrossRefGoogle Scholar
  58. Novel JJ, Nickerson W (1959) Decomposition of native keratin by Streptomyces fradiae. J Bacteriol 77:251–263Google Scholar
  59. Odetallah NH, Wang JJ, Garlich JD, Shih JCH (2003) Keratinase in starter diets improves growth of broiler chicks. Poultry Sci 82:664–670Google Scholar
  60. Onifade AA, Al-Sane NA, Al-Musallam AA, Al-Zarban S (1998) A review: potentials for biotechnological applications of keratin-degrading microorganisms and their enzymes for nutritional improvement of feathers and other keratins as livestock feed resources. Bioresour Technol 66:1–11CrossRefGoogle Scholar
  61. Papadopoulos MC (1986) The effect of enzymatic treatment on amino acid content and nitrogen characteristics of feather meal. Anim Feed Sci Technol 16:151–156CrossRefGoogle Scholar
  62. Pissuwan D, Suntornsuk W (2001) Production of keratinase by Bacillus sp. FK 28 isolated in Thailand. Kasetsart J 35:171–178Google Scholar
  63. Ramnani P, Gupta R (2004) Optimization of medium composition for keratinase production on feather by Bacillus licheniformis RG1 using statistical methods involving response surface methodology. Biotechnol Appl Biochem 40:491–496Google Scholar
  64. Ramnani P, Singh R, Gupta R (2005) Keratinolytic potential of Bacillus licheniformis RG1: structural and biochemical mechanism of feather degradation. Can J Microbiol 51:191–196CrossRefGoogle Scholar
  65. Riffel A, Lucas F, Heeb P, Brandelli A (2003) Characterization of a new keratinolytic bacterium that completely degrades native feather keratin. Arch Microbiol 179:258–265Google Scholar
  66. Rissen S, Antranikian G (2001) Isolation of Thermoanaerobacter keratinophilus sp. nov., a novel thermophilic, anaerobic bacterium with keratinolytic activity. Extremophiles 5:399–408CrossRefGoogle Scholar
  67. Rozs M, Manczinger L, Vagvolgyi C, Kevei F (2001) Secretion of a trypsin-like thiol protease by a new keratinolytic strain of Bacillus licheniformis. FEMS Microbiol Lett 205:221–224CrossRefGoogle Scholar
  68. Sangali S, Brandelli A (2000) Feather keratin hydrolysis by a Vibrio sp. strain kr2. J Appl Microbiol 89:735–743CrossRefGoogle Scholar
  69. Santos RMDB, Firmino AAP, de Sa CM, Felix CR (1996) Keratinolytic activity of Aspergillus fumigatus Fresenius. Curr Microbiol 33:364–370CrossRefGoogle Scholar
  70. Saravanabhavan S, Aravindhan R, Thanikaivelan P, Rao JR, Nair BU, Ramasami T (2004) A source reduction approach: integrated bio-based tanning methods and the role of enzymes in dehairing and fiber opening. Clean Technol Environ Policy 7:3–14CrossRefGoogle Scholar
  71. Schrooyen PMM, Radulf B (2004) Keratin-based products and methods for their productions. US Patent 20,040,210,039Google Scholar
  72. Schrooyen PMM, Dijkstra PJ, Oberthur RC, Bantjes A, Feijen J (2001) Partially carboxymethylated feather keratins. 2. Thermal and mechanical properties of films. J Agric Food Chem 49:221–230CrossRefGoogle Scholar
  73. Singh CJ (1999) Exocellular proteases of Malbranchea gypsea and their role in keratin deterioration. Mycopathologia 143:147–150CrossRefGoogle Scholar
  74. Sohnle PG, Wagner DK (2000) Fungal infections, cutaneous. In: Lederberg J (ed) Encyclopedia of microbiology, 2nd edn. Academic, San Diego, pp 451–459Google Scholar
  75. Stahl ML, Ferrari E (1984) Replacement of the Bacillus subtilis subtilisin structural gene with in-vitro derived mutant. J Bacteriol 158:411–418Google Scholar
  76. Suh HJ, Lee HK (2001) Characterization of a keratinolytic serine protease from Bacillus subtilis KS-1. J Protein Chem 20:165–169CrossRefGoogle Scholar
  77. Suntornsuk W, Suntornsuk L (2003) Feather degradation by Bacillus sp. FK 46 in submerged cultivation. Bioresour Technol 86:239–243CrossRefGoogle Scholar
  78. Szabo L, Benedek A, Szabo ML, Barabas G (2000) Feather degradation with a thermotolerant Streptomyces graminofaciens strain. World J Microbiol Biotechnol 16:252–255CrossRefGoogle Scholar
  79. Takami H, Nogi Y, Horikoshi K (1999) Reidentification of the keratinase-producing facultatively alkaliphilic Bacillus sp. AH-101 as Bacillus halodurans. Extremophiles 3:293–296CrossRefGoogle Scholar
  80. Thanikaivelan P, Rao JR, Nair BU, Ramasami T (2004) Progress and recent trends in biotechnological methods for leather processing. Trends Biotechnol 22:181–188CrossRefGoogle Scholar
  81. Thys RCS, Lucas FS, Riffel A, Heeb P, Brandelli A (2004) Characterization of a protease of a feather-degrading Microbacterium species. Lett Appl Microbiol 39:181–186CrossRefGoogle Scholar
  82. Vidal L, Christen P, Coello MN (2000) Feather degradation by Kocuria rosea in submerged culture. World J Microbiol Biotechnol 16:551–554CrossRefGoogle Scholar
  83. Vignardet C, Guillaume YC, Michel L, Friedrich J, Millet J (2001) Comparison of two hard keratinous substrates submitted to the action of a keratinase using an experimental design. Int J Pharm 224:115–122CrossRefGoogle Scholar
  84. Voet D, Voet JG (1995) Three-dimensional structure of proteins. In: Stiefel J (ed) Biochemistry, 2nd edn. Wiley, New York, pp 154–156Google Scholar
  85. Wang X, Parsons CM (1997) Effect of processing systems on protein quality of feather meal and hair meals. Poultry Sci 76:491–496Google Scholar
  86. Wang JJ, Shih JCH (1999) Fermentation production of keratinase from Bacillus licheniformis PWD-1 and a recombinant B. subtilis FDB-29. J Ind Microbiol Biotechnol 22:608–616CrossRefGoogle Scholar
  87. Wang JJ, Swaisgood HE, Shih JCH (2003a) Bioimmobilization of keratinase using Bacillus subtilis and Escherichia coli systems. Biotechnol Bioeng 81:421–429CrossRefGoogle Scholar
  88. Wang JJ, Swaisgood HE, Shih JCH (2003b) Production and characterization of bio-immobilized keratinase in proteolysis and keratinolysis. Enzyme Microb Technol 32:812–819CrossRefGoogle Scholar
  89. Wang JJ, Rojanatavorn K, Shih JCH (2004) Increased production of Bacillus keratinase by chromosomal integration of multiple copies of the kerA gene. Biotechnol Bioeng 87:459–464CrossRefGoogle Scholar
  90. Wells JA, Ferari E, Henner DJ, Estell DA and Chen EY (1983) Cloning, sequencing and secretion of Bacillus amyloliquefaciens subtilsin in Bacillus subtilis. Nucleic Acids Res 11:7911–7925CrossRefGoogle Scholar
  91. Williams CM, Richter CS, Mackenzie JM Jr, Shih JCH (1990) Isolation, identification and characterization of a feather-degrading bacterium. Appl Environ Microbiol 56:1509–1515Google Scholar
  92. Williams CM, Lee CG, Garlich JD, Shih JCH (1991) Evaluation of a bacterial feather fermentation product, feather-lysate as a feed protein. Poultry Sci 70:85–94Google Scholar
  93. Yamamura S, Morita Y, Hasan Q, Rao SR, Murakami Y, Yokoyama K, Tamiya E (2002a) Characterization of a new keratin-degrading bacterium isolated from deer fur. J Biosci Bioeng 93:595–600Google Scholar
  94. Yamamura S, Morita Y, Hasan Q, Yokoyama K, Tamiya E (2002b) Keratin degradation: a cooperative action of two enzymes from Stenotrophomonas sp. Biochem Biophys Res Commun 294:1138–1143CrossRefGoogle Scholar
  95. Yoshimoto T, Oyama h, Honda T, tone H, Takeshita T, Kamiyama T, Tsuru D (1988) Cloning and expression of subtilisin amylosacchariticus gene. J Biochem 103:1060–1065Google Scholar
  96. Zaghloul TI (1998) Cloned Bacillus subtilis alkaline protease (apr A) gene showing high level of keratinolytic activity. Appl Biochem Biotechnol 70–72:199–205CrossRefGoogle Scholar
  97. Zerdani I, Faid M, Malki A (2004) Feather wastes digestion by new isolated strains Bacillus sp. in Morocco. Afr J Biotechnol 3:67–70Google Scholar

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© Springer-Verlag 2006

Authors and Affiliations

  1. 1.Department of MicrobiologyUniversity of Delhi South CampusNew DelhiIndia

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