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Isolation, Partial purification and Characterization of Keratinase from Bacillus megaterium

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Author Affiliations

  • 1 Dept. of Biochemistry, Center for Post Graduate Studies, Jain University, Jayanagar 3rd Block, Bangalore-560011, Karnataka, INDIA

Int. Res. J. Biological Sci., Volume 2, Issue (2), Pages 13-20, February,10 (2013)

Abstract

A positive keratinase-producing bacterium from soil sample was screened and identified as Bacillus megaterium. The selected growth medium contained feather meal as a sole source of carbon and nitrogen at pH 10.0. The partially purified keratinase has a molecular mass of 41±1 kDa, optimum pH in the alkaline region (9.0) and optimum temperature of 50ºC. and Co2+ slightly inhibited the enzyme activity whereas Hg2+ exhibited a complete inhibition. NEM, IAA, TLCK and NAI moderately inhibited the enzyme whereas PMSF strongly inhibited the keratinase suggesting serine at the active site; however EDTA did not inhibit the enzyme. The isolated strain was thermotolerant and alkalophilic bacterium which can therefore be used in leather processing and detergent industry.

References

  1. Kim J.D., Purification and characterization of a keratinase from a feather-degrading fungus, Aspergillus flavus strain K-03, Microbiol.,35(4), 219-225 (2007)
  2. Brandelli A., Hydrolysis of native proteins by a keratinolytic strain of Chryseobacterium sp. Ann. Microbiol., 55 (1), 47-50 (2005)
  3. Williams C.M., Richter C.S., Mackenzie J.M. and Shih J.C.H., Biocomposites with a content of keratin from chicken feathers, Appl. Environ. Microbiol., 56, 1509-1515 (1990)
  4. Shih J.C.H., Recent development in poultry waste digestion and feather utilization -A review, Poult. Sci.,72, 1617-1620 (1993)
  5. Brutt E.H. and Ichida J. M., Bacteria useful for degrading keratin, United States Patent No. 6, 214,576 B1. 7 p. (2001)
  6. Korkmaz H., Hür H. and Dinçer S., Characterization of alkaline keratinase of Bacillus licheniformis strain HK-1 from poultry waste, Ann. Microbiol., 54(2), 201-211 (2004)
  7. Cai C., Lou B. and Zheng X., Keratinase production and keratin degradation by a mutant strain of Bacillus subtilis, J. Zhejiang Univ. Sci. B, 9(1), 60-67 (2008)
  8. Xu B., Zhong Q., Tang X., Yang Y. and Huang Z., Isolation and characterization of a new keratinolytic bacterium that exhibits significant feather-degrading capability, Afr. J. Biotechnol., 8(18), 4590-4596 (2009)
  9. Gupta R. and Ramnani P., Microbial keratinases and their prospective applications: an overview, Appl. Microbiol. Biotechnol., 70(1), 21-33 (2006)
  10. Mohorcic M., Torkar A., Friedrich J., Kristl J. and Murdan S., An investigation into keratinolytic enzymes to enhance ungual drug delivery, Int. J. Pharm.,332, 196-201 (2007)
  11. Neklyudov A. D., Ivankin A. N. and Berdutina A. V., Properties and uses of protein hydrolysates (review), Appl. Biochem. Microbiol.,36, 452-459 (2000)
  12. Gupta R., Beg Q. and Lorenz P., Bacterial alkaline proteases: molecular approaches and industrial applications, Appl. Microbiol. Biotechnol., 59(1), 15-32 (2002)
  13. Prakash P., Jayalakshmi S. K. and Sreeramulu K., Production of keratinase by free and immobilized cells of Bacillus halodurans strain PPKS-2: partial characterization and its application in feather degradation and dehairing of the goat skin, Appl. Biochem. Biotechnol., 160(7), 1909-1920 (2010)
  14. Zaghloul T.I., Haroun M.A., El-Gayar K. and Abedalal A., Recycling of keratin-containing materials (chicken feather) through genetically engineered bacteria, Polym. Plast. Technol. Eng.43, 1589-1599 (2004)
  15. Caughey B., Interactions between prion protein isoforms: The kiss of death? TIBS, 26, 235-242 (2001)
  16. Tork S., Aly M.M. and Nawar L., Molecular characterization of a new keratinase producing Pseudomomanas sp. MS21, J. Genet. Eng. Biotechnol., 6(1), 37-46 (2008)
  17. Agrahari S. and Wadhwa N., Degradation of chicken feather a poultry waste product by keratinolytic bacteria isolated from dumping site at Ghazipur poultry processing plant, Int. J. Poult. Sci.,9(5), 482-489 (2010)
  18. Sneath P.H.A., Mair N.S., Sharpe M.E. and Holt J.G., Bergey’s Manual of Systematic Bacteriology, Vol. 2 Baltimore., Williams and Wilkins (1986)
  19. Lin H.H. and Yin L.J., Feather meal and rice husk enhanced keratinase production by Bacillus licheniformis YJ4 and characterization of produced keratinases, J. Mar. Sci. Technol., 18(3), 458-465 (2010)
  20. Wawrzkiewicz K., Lobarzewski J. and Wolski T., Intracellular keratinase of Trichophytongallinae,Med. Mycol.25(4), 261-268 (1987)
  21. Vigneshwaran C., Shanmugam S. and Kumar T. S., Screening and characterization of keratinase from Bacillus licheniformis isolated from Namakkal poultry farm, Researcher, 2(4), 89-96 (2010)
  22. Lowry O.H., Rosebrough N.J., Farr A.L. and Randall R.J., Protein measurement with the Folin phenol reagent, J. Biol. Chem.,193, 265-275 (1951)
  23. Ali T. H., Ali N. H. and Mohamed L. A., Production, purification and some properties of extracellular keratinase from feathers-degradation by Aspergillus oryzae NRLL-447. J. Appl. Sci. Environ. Sanit., 6 (2), 123-136 (2011)
  24. Laemmli U.K., Cleavage of structural proteins during the assembly of the head of bacteriophage T4, Nature, 227(5259) 680-685 (1970)
  25. Blum H., Beier H. and Gross H.J., Improved silver staining of plant proteins, RNA and DNA in polyacrylamide gels, Electrophoresis, 8(2), 93-99 (1987)
  26. Brandelli A., Daroit D. J. and Riffel A., Biochemical features of microbial keratinases and their production and applications, Appl. Microbiol. Biotechnol. 85(6), 1735-1750 (2010)
  27. Wood M., Environmentasl oil biology, Blackie Academic and Professional, London (1995)
  28. Cheng S.W., Hu M.N., Shen S.W., Takagi H., Asano M. and Tsai Y.C., Production and characterization of keratinase of a feather-degrading Bacillus licheniformisPWD-1, Biosci. Biotech. Biochem., 59, 2239-2243 (1995)
  29. Kainoor P.S. and Naik G.R., Production and charecterization of feather degrading keratinase from Bacillus sp. JB 99, Indian J. Biotechnol., 384-390 (2010)
  30. Srivastava A., Sharma A. and Suneetha V., Feather waste biodegradation as a source of amino acid, Eur. J. Exp. Biol., 1(2), 56-63 (2011)
  31. Brandelli A. and Riffel A., Production of an extracellular keratinase from Chryseobacterium sp. growing on raw feathers, Electr. J. Biotechnol., 8(1), 35-42 (2005)
  32. Suh H. J., Kim J. M. and Lim W. J., Feather-degrading Bacillus species from poultry waste, Process Biochem., 37, 287-291 (2001)
  33. Suntornsuk W., Tongjun J., Onnim P., Oyama H., Ratanakanokchai K., Kusamran T. and Oda K., Purification and characterisation of keratinase from a thermotolerant feather-degrading bacterium, World J. Microbiol. Biotechnol., 21, 1111-1117 (2004)
  34. Dozie I.N.S., Okeke C.N. and Unaeze N.C., A thermostable alkaline active keratinolytic proteinase from Crysosporium keratinophylum, Word J. Microbiol. Biotechnol., 10, 563-567 (1994)
  35. Bressollier P., Letourneau F., Urdaci M. and Verneuil B., Purification and characterization of a keratinolytic serine proteinase from Streptomyces albidoflavus, Appl. Environ. Microbiol., 65(6), 2570-2576 (1999)
  36. Bernal C., Cairo J. and Coello N., Purification and characterization of a novel exocellular keratinase from Kocuria rosea. Enzyme Microbial. Technol., 38, 49-54 (2006)
  37. Chitte R.R., Nalawade V.K. and Dey S., Keratinolytic activity from the broth of a feather-degrading thermophilic Streptomyces thermoviolaceus strain SD8, Lett. Appl. Microbiol., 28, 131-136 (1999)
  38. Kumar A.G., Swarnalatha S., Gayathri S., Nagesh N. and Sekaran G., Characterization of an alkaline active - thiol forming extracellular serine keratinase by the newly isolated Bacillus pumilus, J. Appl. Microbiol., 104, 411-419 (2007)
  39. Rozs M., Manczinger L., Vagvlgyi C. and Kevei F., Secretion of a trypsin-like thiol protease by a new keratinolytic strain of Bacillus licheniformis, FEMS Microbiol. Lett.,205, 221-224 (2001)
  40. Thys R.C.S., Lucas F.S., Riffel A., Heeb P. and Brandelli A., Characterization of a protease of a feather-degrading Microbacterium species, Lett. Appl. Microbiol., 39(2), 181-186 (2004)
  41. Lusrerio D.D., Suizo F.G., Labunos N.M., Valledor M.N.,Ueda S., Kawai A.S., Koike K. and Shikata S., Alkaliresistant endo-1 4-ß-gluconase produce by Bacillus sp.PKM-5430, Biosci. Biotechnol. Biochem., 56, 1671-1672(1992)