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High Keratinase Production And Keratin Degradation By A Mutant Strain Kr II, Derived From Streptomyces radiopugnans Kr I2

Year 2018, Volume: 12 Issue: 2, 1 - 7, 25.09.2018

Abstract

This study aimed to purify and characterize a new feather-degrading enzyme. Twenty bacterial isolates were recovered from different sources on feather-meal medium. The best keratinase producer KR12, isolated from a poultry farm, was selected for the further experiments. Physiological and biochemical studies indicated that the bacterium KR12 belongs to genus Streptomyces. 16S rRNA analysis confirmed this result since the KR12 was similar by 98.8% to Streptomyces radiopugnans. The enzyme was purified using Sephadex G-75 and DEAE (diethylaminoethyl)-Sepharose chromatographic columns. The purified enzyme had a molecular weight of 32 kDa, defined by sodium dodecyl sulfate polyacrylamide gel electrophoresis analysis (SDS-PAGE). The enzyme was active between 30 and 60°C. The enzymatic activity of the purified enzyme was enhanced by Zn2+, Na+, Ca2+, K+, and inhibited by Fe2+, Cu2+, and EDTA, demonstrating that keratinase from Streptomyces radiopugnans KR 12 belongs to metallo keratinases. Keratinase production was enhanced using UV radiation. Three recombinant mutants were obtained, mutant KR II, produced a high keratinase enzyme (3 fold) than the wild strain. Streptomyces radiopugnans KR12 and its mutant are promising for keratinase production and many biotechnological applications.

References

  • Onifade, AA, A1-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.
  • Gupta R, Ramnani.P. 2006. Microbial keratinase, and their prospective application: an overview. Appl. Microbiol. Biotechnol., 70:21-33.
  • Lin X, Shih JCH and Swaisgood HE. 1996. Hydrolysis of feather keratin by immobilized keratinase. Appl. Environ. Microbiol. 62:4273-4275.
  • Friedrich J, Gradisar H, Mandin D and Chaumont JP. 1999. Screening fungi for the synthesis of keratinolytic enzymes. Lett. Applied Microbiol. 28: 127-130.
  • Essien JP, Umoh AA, Akpan EJ, Eduok SI and Umoiyoho A. 2009. Growth, keratinolytic proteinase activity and thermotolerance of dermatophytes associated with alopecia in Uyo, Nigeria. Acta Microbiol. Et Immunol. Hungarica, 56: 61-69.
  • Anbu P, Hilda A, Sur H, Hur B and Jayanthi S. 2008. Extracellular keratinase from Trichophyton sp. HA-2 isolated from feather dumping soil. Intern. Biodeterior. Biodegrad. 62: 287-292.
  • Cai C and Zheng X. 2009. Medium optimization for keratinase production in hair substrate by a new Bacillus subtilis KD-N2 using response surface methodology. J. Ind. Microbiol. Biotechnol., 36: 875-883.
  • Macedo AJ, da Silva WOB, Gava R, Driemeier D, Henriques JP, Termignoni C. 2005.Novel keratinase from Bacillus subtilis S14 exhibiting remarkable dehairing capabilities. Appl. Environ. Microbiol.71: 594-596.
  • Pillai P and Archana G. 2008. Hide depilation and feather disintegration studies with keratinolytic serine protease from a novel Bacillus subtilis isolate. Applied Microbiol. Biotechnol. 78: 643-650.
  • Tork S, Shahin Y, El-Hakim A, Abdel-Aty A, Aly M. 2016. Novel Extracellular Bacillus pumilus NRC21 serine Metallo-keratinase with potential keratin degrading capacity. Int. j. Biol. Macromol. 86: 189-196.
  • Tatineni R, Doddapanem KK, Potumarthi RC, Vellanki RN, Kandathil, MT, Kolli N and Mangamoori. LN. 2008. Purification and characterization of an alkaline keratinase from Streptomyces sp. Bioresour. Technol. 99: 1596-1602.
  • M. M. Aly and S. Tork / JABS, 12 (2): 01-07, 2018 7
  • Syed DG, Lee JC, Li WJ, Kim CJ and Agasar D. 2009. Production, characterization and application of keratinase from Streptomyces gulbargensis. Bioresour. Technol. 100: 1868-1871.
  • Horneck G, Bücker H, Reitz G, Requardt H, Dose K, Martens KD, Mennigmann HD and Weber P. 1984. Microorganisms in the space environment. Science. 225:226-228.
  • Apichaisataienchote B, AltenbuchnerJ and Buchenauer H.2005. Isolation and Identification of Streptomyces fradiae SU-1 from Thailand and Protoplast Transformation with the Chitinase B Gene from Nocardiopsisprasina OPC-131, CurrMicrobiol. 51: 116-121,
  • Saran S, Isar J and Saxena RK. 2007.A modified method for the detection of microbial proteases on agar plates using tannic acid. J. Biochem. Biophysical Methods. 70: 697-699.
  • Sambrook J and Russell DW. Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA, 3rd edition. 2001.
  • Shirling EB and GottliebD. 1966. Methods for characterization of Streptomyces species. Int. J. Syst. Bacteriol. 16: 313-340.
  • Williams ST, Sharpe ME and Molt JC. 1989. In: Bergey's Manual of Systematic Bacteriology (Williams and Wilkins (eds). 4: 2300-2648.
  • Agwa A, Aly MM and Bonaly R. 2000. Isolation and characterization of two Streptomyces species produced nonpolyenic antifungal agents. J. Union Arab Biol. 7: 62- 84.
  • Hasegawa T, Takizawa M .and Tanida S. 1983. A rapid analysis for chemical grouping of aerobic actinomycetes. J Gen Appl Microbiol .29: 319–322.
  • Butte W. 1983. Rapid method for the determination of fatty acid profiles from fats and oils using trimethylsulphonium hydroxide for transesterification. J. Chromatography. 261: 142-1 45.
  • Hoischen C, Ihn W, Gura K and Gumpert J. 1997. Structural characterization of molecular phospholipid species in cytoplasmic membranes of the cell wall-less Streptomyces hygroscopicus L form by use of electrospray ionization coupled with collision-induced dissociation mass spectrometry. J. Bacteriol. 179: 3437-3442.
  • Weisberg WG, Barns SM, Pelletier DA and Lane DJ. 1991.16S Ribosomal DNA Amplification for Phylogenetic Study. J. Bacteriol. 173: 697-703.
  • Tork S, Aly MM, Nawar L. 2010. Biochemical and molecular characterization of a new local keratinase producing Pseudomonas sp., MS21. Asian J. Biotechnol. 2: 1-13.
  • Letourneau F, Soussotte P, Bressollier P, Branland P and Verneuil B. 1982. Keratinolytic activity of Streptomyces sp. SK1-02 a new isolated strain. Lett. Applied Microbiol. 26: 77-80.
  • Scott JA and Untereinen.WA. 2004.Determination of keratin degradation by fungi using keratin azure. Med. Mycol. 42: 239-246.
  • Bradford MM. 1976.A rapid and sensitive for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72: 248-254.
  • Laemmli U.K. 1970. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 227: 680–685.
  • Sangali S and Brandelli A. 2000. Feather keratin hydrolysis by a Vibrio sp. strain kr2. J. Applied Microbiol. 89: 735-743.
  • Doi E, Shibata D, Matoba T. 1981. Modified colorimetric ninhydrin methods for peptidase. assay. Anal. Biochem. 118: 173–184.
  • Suntornsuk W, Tongjun J, Onnim P, Oyama H, Ratanakanokchai K, Kusamran T, Oda K. 2005. Purification and characterization of keratinase from a thermotolerant feather-degrading bacterium. J. Ind. Microbiol. Biotechnol. 21: 1111-1117.
  • Chitte RR, Nalawade V K and Dey S. 1999. Keratinolytic activity from the broth of a feather-degrading thermophilic Streptomyces thermoviolaceus strain SD8. Let. Appl. Microbiol. 28: 131–136.
  • Nam GW, Lee DW, Lee HS, Lee NJ, Kim B.C, Choe, EA, Hwang JK, Suhartono MT, Pyun.YR. 2002. Native-feather degradation by Fervidobacteriumislandicum AW-1, a newly isolated keratinase-producing thermophilic anaerobe. Arch Microbiol. 178: 538- 547.
  • Bernal C, Cairo J, Coello N. 2006. Purification and characterization of a novel exocellular keratinase from Kocuriarosea. Enzyme Microb. Technol. 38: 49-54.
  • Bockle B, Galunski, B, Müller R. 1995. Characterization of a keratinolytic serine protease from Streptomyces pactum DSM40530. Appl. Environ. Microbiol. 61: 3705-3710.
  • Gessesse A, Hatti-Kaul R, Gashe BA, Mattiasson B. 2003. Novel alkaline proteases from alkaliphilic bacteria grown on the chicken feather. Enzyme Microb. Technol., 32:519-524.
  • Gradišar H, Friedrich J, Križaj I, Jerala R. 2005.Similarities and specificities of fungal keratinolytic proteases: comparison of keratinase of Paecilomyces marquandii and Doratomyces microsporus to some known proteases. Appl Environ Microbiol. 71:3420–3426.
  • Tan TW, Zhang M, Wang BW, Ying CH and Deng. 2003. Screening of high lipase producing Candida sp. and production of lipase by fermentation. Process Biochem. 39:459–465.
  • Wang CL, Li DF, Lu WQ, Wang YH and Lai CH. 2004. Influence of cultivating conditions on the alpha-galactosidase biosynthesis from a novel strain of Penicillium sp. in solid-state fermentation. Lett Appl Microbiol. 39:369–375.
  • Cai CG, Lou BG, Zheng XD. 2008. Keratinase production and keratin degradation by a mutant strain of Bacillus subtilis. J Zhejiang Univ Sci B. 9: 60–67.
Year 2018, Volume: 12 Issue: 2, 1 - 7, 25.09.2018

Abstract

References

  • Onifade, AA, A1-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.
  • Gupta R, Ramnani.P. 2006. Microbial keratinase, and their prospective application: an overview. Appl. Microbiol. Biotechnol., 70:21-33.
  • Lin X, Shih JCH and Swaisgood HE. 1996. Hydrolysis of feather keratin by immobilized keratinase. Appl. Environ. Microbiol. 62:4273-4275.
  • Friedrich J, Gradisar H, Mandin D and Chaumont JP. 1999. Screening fungi for the synthesis of keratinolytic enzymes. Lett. Applied Microbiol. 28: 127-130.
  • Essien JP, Umoh AA, Akpan EJ, Eduok SI and Umoiyoho A. 2009. Growth, keratinolytic proteinase activity and thermotolerance of dermatophytes associated with alopecia in Uyo, Nigeria. Acta Microbiol. Et Immunol. Hungarica, 56: 61-69.
  • Anbu P, Hilda A, Sur H, Hur B and Jayanthi S. 2008. Extracellular keratinase from Trichophyton sp. HA-2 isolated from feather dumping soil. Intern. Biodeterior. Biodegrad. 62: 287-292.
  • Cai C and Zheng X. 2009. Medium optimization for keratinase production in hair substrate by a new Bacillus subtilis KD-N2 using response surface methodology. J. Ind. Microbiol. Biotechnol., 36: 875-883.
  • Macedo AJ, da Silva WOB, Gava R, Driemeier D, Henriques JP, Termignoni C. 2005.Novel keratinase from Bacillus subtilis S14 exhibiting remarkable dehairing capabilities. Appl. Environ. Microbiol.71: 594-596.
  • Pillai P and Archana G. 2008. Hide depilation and feather disintegration studies with keratinolytic serine protease from a novel Bacillus subtilis isolate. Applied Microbiol. Biotechnol. 78: 643-650.
  • Tork S, Shahin Y, El-Hakim A, Abdel-Aty A, Aly M. 2016. Novel Extracellular Bacillus pumilus NRC21 serine Metallo-keratinase with potential keratin degrading capacity. Int. j. Biol. Macromol. 86: 189-196.
  • Tatineni R, Doddapanem KK, Potumarthi RC, Vellanki RN, Kandathil, MT, Kolli N and Mangamoori. LN. 2008. Purification and characterization of an alkaline keratinase from Streptomyces sp. Bioresour. Technol. 99: 1596-1602.
  • M. M. Aly and S. Tork / JABS, 12 (2): 01-07, 2018 7
  • Syed DG, Lee JC, Li WJ, Kim CJ and Agasar D. 2009. Production, characterization and application of keratinase from Streptomyces gulbargensis. Bioresour. Technol. 100: 1868-1871.
  • Horneck G, Bücker H, Reitz G, Requardt H, Dose K, Martens KD, Mennigmann HD and Weber P. 1984. Microorganisms in the space environment. Science. 225:226-228.
  • Apichaisataienchote B, AltenbuchnerJ and Buchenauer H.2005. Isolation and Identification of Streptomyces fradiae SU-1 from Thailand and Protoplast Transformation with the Chitinase B Gene from Nocardiopsisprasina OPC-131, CurrMicrobiol. 51: 116-121,
  • Saran S, Isar J and Saxena RK. 2007.A modified method for the detection of microbial proteases on agar plates using tannic acid. J. Biochem. Biophysical Methods. 70: 697-699.
  • Sambrook J and Russell DW. Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA, 3rd edition. 2001.
  • Shirling EB and GottliebD. 1966. Methods for characterization of Streptomyces species. Int. J. Syst. Bacteriol. 16: 313-340.
  • Williams ST, Sharpe ME and Molt JC. 1989. In: Bergey's Manual of Systematic Bacteriology (Williams and Wilkins (eds). 4: 2300-2648.
  • Agwa A, Aly MM and Bonaly R. 2000. Isolation and characterization of two Streptomyces species produced nonpolyenic antifungal agents. J. Union Arab Biol. 7: 62- 84.
  • Hasegawa T, Takizawa M .and Tanida S. 1983. A rapid analysis for chemical grouping of aerobic actinomycetes. J Gen Appl Microbiol .29: 319–322.
  • Butte W. 1983. Rapid method for the determination of fatty acid profiles from fats and oils using trimethylsulphonium hydroxide for transesterification. J. Chromatography. 261: 142-1 45.
  • Hoischen C, Ihn W, Gura K and Gumpert J. 1997. Structural characterization of molecular phospholipid species in cytoplasmic membranes of the cell wall-less Streptomyces hygroscopicus L form by use of electrospray ionization coupled with collision-induced dissociation mass spectrometry. J. Bacteriol. 179: 3437-3442.
  • Weisberg WG, Barns SM, Pelletier DA and Lane DJ. 1991.16S Ribosomal DNA Amplification for Phylogenetic Study. J. Bacteriol. 173: 697-703.
  • Tork S, Aly MM, Nawar L. 2010. Biochemical and molecular characterization of a new local keratinase producing Pseudomonas sp., MS21. Asian J. Biotechnol. 2: 1-13.
  • Letourneau F, Soussotte P, Bressollier P, Branland P and Verneuil B. 1982. Keratinolytic activity of Streptomyces sp. SK1-02 a new isolated strain. Lett. Applied Microbiol. 26: 77-80.
  • Scott JA and Untereinen.WA. 2004.Determination of keratin degradation by fungi using keratin azure. Med. Mycol. 42: 239-246.
  • Bradford MM. 1976.A rapid and sensitive for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72: 248-254.
  • Laemmli U.K. 1970. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 227: 680–685.
  • Sangali S and Brandelli A. 2000. Feather keratin hydrolysis by a Vibrio sp. strain kr2. J. Applied Microbiol. 89: 735-743.
  • Doi E, Shibata D, Matoba T. 1981. Modified colorimetric ninhydrin methods for peptidase. assay. Anal. Biochem. 118: 173–184.
  • Suntornsuk W, Tongjun J, Onnim P, Oyama H, Ratanakanokchai K, Kusamran T, Oda K. 2005. Purification and characterization of keratinase from a thermotolerant feather-degrading bacterium. J. Ind. Microbiol. Biotechnol. 21: 1111-1117.
  • Chitte RR, Nalawade V K and Dey S. 1999. Keratinolytic activity from the broth of a feather-degrading thermophilic Streptomyces thermoviolaceus strain SD8. Let. Appl. Microbiol. 28: 131–136.
  • Nam GW, Lee DW, Lee HS, Lee NJ, Kim B.C, Choe, EA, Hwang JK, Suhartono MT, Pyun.YR. 2002. Native-feather degradation by Fervidobacteriumislandicum AW-1, a newly isolated keratinase-producing thermophilic anaerobe. Arch Microbiol. 178: 538- 547.
  • Bernal C, Cairo J, Coello N. 2006. Purification and characterization of a novel exocellular keratinase from Kocuriarosea. Enzyme Microb. Technol. 38: 49-54.
  • Bockle B, Galunski, B, Müller R. 1995. Characterization of a keratinolytic serine protease from Streptomyces pactum DSM40530. Appl. Environ. Microbiol. 61: 3705-3710.
  • Gessesse A, Hatti-Kaul R, Gashe BA, Mattiasson B. 2003. Novel alkaline proteases from alkaliphilic bacteria grown on the chicken feather. Enzyme Microb. Technol., 32:519-524.
  • Gradišar H, Friedrich J, Križaj I, Jerala R. 2005.Similarities and specificities of fungal keratinolytic proteases: comparison of keratinase of Paecilomyces marquandii and Doratomyces microsporus to some known proteases. Appl Environ Microbiol. 71:3420–3426.
  • Tan TW, Zhang M, Wang BW, Ying CH and Deng. 2003. Screening of high lipase producing Candida sp. and production of lipase by fermentation. Process Biochem. 39:459–465.
  • Wang CL, Li DF, Lu WQ, Wang YH and Lai CH. 2004. Influence of cultivating conditions on the alpha-galactosidase biosynthesis from a novel strain of Penicillium sp. in solid-state fermentation. Lett Appl Microbiol. 39:369–375.
  • Cai CG, Lou BG, Zheng XD. 2008. Keratinase production and keratin degradation by a mutant strain of Bacillus subtilis. J Zhejiang Univ Sci B. 9: 60–67.
There are 41 citations in total.

Details

Primary Language English
Journal Section Research Article
Authors

Magda M. Aly This is me

Sanaa Tork

Publication Date September 25, 2018
Published in Issue Year 2018 Volume: 12 Issue: 2

Cite

APA Aly, M. M., & Tork, S. (2018). High Keratinase Production And Keratin Degradation By A Mutant Strain Kr II, Derived From Streptomyces radiopugnans Kr I2. Journal of Applied Biological Sciences, 12(2), 1-7.
AMA Aly MM, Tork S. High Keratinase Production And Keratin Degradation By A Mutant Strain Kr II, Derived From Streptomyces radiopugnans Kr I2. J.appl.biol.sci. September 2018;12(2):1-7.
Chicago Aly, Magda M., and Sanaa Tork. “High Keratinase Production And Keratin Degradation By A Mutant Strain Kr II, Derived From Streptomyces Radiopugnans Kr I2”. Journal of Applied Biological Sciences 12, no. 2 (September 2018): 1-7.
EndNote Aly MM, Tork S (September 1, 2018) High Keratinase Production And Keratin Degradation By A Mutant Strain Kr II, Derived From Streptomyces radiopugnans Kr I2. Journal of Applied Biological Sciences 12 2 1–7.
IEEE M. M. Aly and S. Tork, “High Keratinase Production And Keratin Degradation By A Mutant Strain Kr II, Derived From Streptomyces radiopugnans Kr I2”, J.appl.biol.sci., vol. 12, no. 2, pp. 1–7, 2018.
ISNAD Aly, Magda M. - Tork, Sanaa. “High Keratinase Production And Keratin Degradation By A Mutant Strain Kr II, Derived From Streptomyces Radiopugnans Kr I2”. Journal of Applied Biological Sciences 12/2 (September 2018), 1-7.
JAMA Aly MM, Tork S. High Keratinase Production And Keratin Degradation By A Mutant Strain Kr II, Derived From Streptomyces radiopugnans Kr I2. J.appl.biol.sci. 2018;12:1–7.
MLA Aly, Magda M. and Sanaa Tork. “High Keratinase Production And Keratin Degradation By A Mutant Strain Kr II, Derived From Streptomyces Radiopugnans Kr I2”. Journal of Applied Biological Sciences, vol. 12, no. 2, 2018, pp. 1-7.
Vancouver Aly MM, Tork S. High Keratinase Production And Keratin Degradation By A Mutant Strain Kr II, Derived From Streptomyces radiopugnans Kr I2. J.appl.biol.sci. 2018;12(2):1-7.