Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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Study of the Production of Alkaline Keratinases in Submerged Cultures as an Alternative for Solid Waste Treatment Generated in Leather Technology

  • Cavello, Ivana A. (Research and Development Center for Industrial Fermentations (CINDEFI, UNLP: CCT-La Plata, CONICET)) ;
  • Chesini, Mariana (Research and Development Center for Industrial Fermentations (CINDEFI, UNLP: CCT-La Plata, CONICET)) ;
  • Hours, Roque A. (Research and Development Center for Industrial Fermentations (CINDEFI, UNLP: CCT-La Plata, CONICET)) ;
  • Cavalitto, Sebastian F. (Research and Development Center for Industrial Fermentations (CINDEFI, UNLP: CCT-La Plata, CONICET))
  • Received : 2012.11.07
  • Accepted : 2013.04.04
  • Published : 2013.07.28
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Abstract

Six nonpathogenic fungal strains isolated from alkaline soils of Buenos Aires Province, Argentina (Acremonium murorum, Aspergillus sidowii, Cladosporium cladosporoides, Neurospora tetrasperma, Purpureocillium lilacinum (formerly Paecilomyces lilacinus), and Westerdikella dispersa) were tested for their ability to produce keratinolytic enzymes. Strains were grown on feather meal agar as well as in solid-state and submerged cultures, using a basal mineral medium and "hair waste" as sole sources of carbon and nitrogen. All the tested fungi grew on feather meal agar, but only three of them were capable of hydrolyzing keratin, producing clear zones. Among these strains, P. lilacinum produced the highest proteolytic and keratinolytic activities, both in solid-state and submerged fermentations. The medium composition and culture conditions for the keratinases production by P. lilacinum were optimized. Addition of glucose (5 g/l) and yeast extract (2.23 g/l) to the basal hair medium increased keratinases production. The optimum temperature and initial pH for the enzyme production were $28^{\circ}C$ and 6.0, respectively. A beneficial effect was observed when the original concentration of four metal ions, present in the basal mineral medium, was reduced up to 1:10. The maximum yield of the enzyme was 15.96 $U_c/ml$ in the optimal hair medium; this value was about 6.5-fold higher than the yield in the basal hair medium. These results suggest that keratinases from P. lilacinum can be useful for biotechnological purposes such as biodegradation (or bioconversion) of hair waste, leading to a reduction of the environmental pollution caused by leather technology with the concomitant production of proteolytic enzymes and protein hydrolyzates.

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References

  1. Agrebi R, Haddar A, Hajji M, Frikha F, Manni L, Jellouli K. 2009. Fibrinolytic enzymes from a newly isolated marine bacterium Bacillus subtilis A26: Characterization and statistical media optimization. Can. J. Microbiol. 55: 1049-1061. https://doi.org/10.1139/W09-057
  2. Allpress JD, Mountain G, Gowland PC. 2002. Production, purification and characterization of an extracellular keratinase from Lysobacter NCIMB 9407. Lett. Appl. Microbiol. 34: 337-342. https://doi.org/10.1046/j.1472-765X.2002.01093.x
  3. Anbu P, Gopinath SCB, Hilda A, Lakshmipriya T, Annadurai G. 2005. Extracellular keratinase from Trychophyton sp. HA-2 isolated from leather dumping soil. Enzyme Microb. Technol. 36: 639-647. https://doi.org/10.1016/j.enzmictec.2004.07.019
  4. Arrondo JL, Young NM, Mantsch HH. 1988. The solution structure of concanavalin A probed by FT-IR spectroscopy. Biochim. Biophys. Acta 952: 261-268. https://doi.org/10.1016/0167-4838(88)90125-2
  5. Balint B, Bagi Z, Toth A, Rakhely G, Perei K, Kovacs K. 2005. Utilization of keratin-containing biowaste to produce biohydrogen. Appl. Microbiol. Biotechnol. 69: 404-410. https://doi.org/10.1007/s00253-005-1993-3
  6. Banerjee R, Bhattacharya BC. 1992. Extracellular alkaline protease of newly isolated Rhizopus oryzae. Biotechnol. Lett. 14: 301-304. https://doi.org/10.1007/BF01022328
  7. Bradford MM. 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72: 248-254. https://doi.org/10.1016/0003-2697(76)90527-3
  8. Brandelli A. 2005. Hydrolysis of native proteins by a keratinolytic protease of Chryseobacterium sp. Ann. Microbiol. 55: 47-50.
  9. Chon D, Kwon T. 2001. Isolation of keratinolytic protease producing microorganism and its cultivation condition. San'oeb Misaengmul Haghoeji 29: 134-141.
  10. Correa AP, Daroit DJ, Brandelli A. 2010. Characterization of a keratinase produced by Bacillus sp. P7 isolated from an Amazonian environment. Int. Biodeter. Biodegrad. 64: 1-6. https://doi.org/10.1016/j.ibiod.2009.06.015
  11. Daroit DJ, Simonetti A, Hertz PF, Brandelli A. 2008. Purification and characterization of an extracellular betaglucosidase from Monascus purpureus. J. Microbiol. Biotechnol. 18: 933-941.
  12. De Azaredo LAI, De Lima MB, Coelho RRR, Freire DM. 2006. Thermophilic protease production by Streptomyces sp. 594 in submerged and solid-state fermentations using feather meal. J. Appl. Microbiol. 100: 641-647. https://doi.org/10.1111/j.1365-2672.2005.02791.x
  13. El-Gendy MMA. 2010. Keratinase production by endophytic Penicillium spp. Morsy1 under solid-state fermentation using rice straw. Appl. Biochem. Biotechnol. 162: 780-794. https://doi.org/10.1007/s12010-009-8802-x
  14. El-Refai HA, AbdelNaby MA, Gaballa A, El-Araby MH, Abdel Fattah AF. 2005. Improvement of the newly isolated Bacillus pumilus FH9 keratinolytic activity. Process Biochem. 40: 2325-2332. https://doi.org/10.1016/j.procbio.2004.09.006
  15. Eliades L, Cabello M, Voget CE, Galarza B, Saparrat M. 2010. Screening for alkaline keratinolytic activity in fungi isolated from soils of the biosphere reserve "Parque Costero del Sur" (Argentina). World J. Microbiol. Biotechnol. 26: 2105-2111. https://doi.org/10.1007/s11274-010-0389-4
  16. Farag AM, Hassan MA. 2004. Purification, characterization and immobilization of a keratinase from Aspergillus oryzae. Enzyme Microb. Technol. 34: 85-93. https://doi.org/10.1016/j.enzmictec.2003.09.002
  17. Ferreyra OA, Cavalitto SF, Hours RA, Ertola RJ. 2002. Influence of trace elements on enzyme production: Protopectinase expression by a Geotrichum klebahnii strain. Enzyme Microb. Technol. 31: 498-504. https://doi.org/10.1016/S0141-0229(02)00146-1
  18. Friedrich AB, Antranikian G. 1996. Keratin degradation by Fervidobacterium pennovorans, a novel thermophilic anaerobic species of the order Thermotogales. Appl. Environ. Microbiol. 62: 2875-2882.
  19. Galarza B, Goya L, Cantera C, Garro ML, Reinoso HM, Lopez LMI. 2004. Fungal biotransformation of bovine hair part I: Isolation of fungus with keratinolytic activity. J. Soc. Leather Technol. Chem. 88: 93-98.
  20. Galarza BC, Goya L, Garro ML, Mercerat J, Hours RA, Cantera CS. 2005. Fungal biotransformation of bovine hair part II: Biomass and proteases produced as a function of incubation time. Assessment of hair waste digestion. J. Soc. Leather Technol. Chem. 90: 169-172.
  21. Garcia-Carreno FL, Dimes LE, Haard NF. 1993. Substrategel electrophoresis for composition and molecular weight of proteinases or proteinaceous proteinase inhibitors. Anal. Biochem. 214: 65-69. https://doi.org/10.1006/abio.1993.1457
  22. Gradisar H, Kern S, Friedrich J. 2000. Keratinase of Doratomyces microsporus. Appl. Microbiol. Biotechnol. 53: 196-200. https://doi.org/10.1007/s002530050008
  23. Gupta R, K Beg, P Lorenz. 2002. Bacterial alkaline proteases: Molecular approaches and industrial applications. Appl. Microbiol. Biotechnol. 59: 15-32. https://doi.org/10.1007/s00253-002-0975-y
  24. Hossain MS, Azad AK, Abu Sayem SM, Mostafa G, Mozammel Hoq Md. 2007. Production and partial characterization of feather-degrading keratinolytic serine protease from Bacillus licheniformis MZK-3. J. Biol. Sci. 7: 599-606. https://doi.org/10.3923/jbs.2007.599.606
  25. Huang Q, Peng Y, Li X, Wang H, Zhang Y. 2003. Purification and characterization of an extracellular alkaline serine protease with dehairing function from Bacillus pumilus. Curr. Microbiol. 46: 169-173. https://doi.org/10.1007/s00284-002-3850-2
  26. Jeong J, Lee O, Jeon Y, Kim J, Lee N. 2010. Production of keratinolytic enzyme by a newly isolated feather-degrading Stenotrophomonas maltophilia that produces plant growthpromoting activity. Process Biochem. 45: 1738-1745. https://doi.org/10.1016/j.procbio.2010.07.020
  27. Joshi SG, Tejashwini MM, Revati N, Sridevi R, Roma D. 2007. Isolation, identification and characterization of a feather degrading bacterium. Int. J. Poultry Sci. 6: 689-693. https://doi.org/10.3923/ijps.2007.689.693
  28. Kainoor P, Naik GR. 2010. Production and characterization of feather degrading keratinase from Bacillus sp. JB 99. Indian J. Biotechnol. 9: 384-390.
  29. Kumar R, Balaji S, Uma TS, Mandal AB, Sehgal PK. 2010. Optimization of influential parameters for extracellular keratinase production by Bacillus subtilis (MTCC9102) in solid state fermentation using horn meal - a biowaste management. Appl. Biochem. Biotechnol. 160: 30-39. https://doi.org/10.1007/s12010-008-8452-4
  30. Laemmli UK. 1970. Cleavage of structural proteins during assembly of head of bacteriophage T4. Nature 227: 680-685. https://doi.org/10.1038/227680a0
  31. Lee Y, Kim J, Kim H, Lee J. 2004. Production and characterization of keratinase from Paracoccus sp. WJ-98. Biotechnol. Bioproc. E 9: 17-22. https://doi.org/10.1007/BF02949317
  32. Liggieri C, Arribere MC, Trejo S, Canals F, Aviles F, Priolo N. 2004. Purification and biochemical characterization of asclepain c I from the latex of Asclepias curassavica L. Protein J. 23: 403-411. https://doi.org/10.1023/B:JOPC.0000039554.18157.69
  33. Macedo AJ, Beys da Silva WO, Gava R, Driemeier D, Pegas Henriques JA, Termignoni C. 2005. Novel keratinase from Bacillus subtilis S14 exhibiting remarkable dehairing capabilities. Appl. Environ. Microbiol. 71: 594-596. https://doi.org/10.1128/AEM.71.1.594-596.2005
  34. Marcondes NR, Taira CL, Vandresen DC, Svidzinski TIE, Kadowaki MK, Peralta RM. 2008. New feather-degrading filamentous fungi. Microb. Ecol. 56: 13-17. https://doi.org/10.1007/s00248-007-9319-x
  35. Mazotto AM, Lage Cedrola SM, Lins U, Rosado AS, Silva KT, Chaves JQ, et al. 2010. Keratinolytic activity of Bacillus subtilis AMR using human hair. Lett. Appl. Microbiol. 50: 89-96. https://doi.org/10.1111/j.1472-765X.2009.02760.x
  36. Moallaei H, Zaini F, Larcher G, Beucher B, Bouchara JP. 2006. Partial purification and characterization of a 37 KDa extracellular proteinase from Trichophyton vanbreuseghemii. Mycopathologia 161: 369-375. https://doi.org/10.1007/s11046-006-0019-8
  37. Muga A, Arrondo JL, Bellon T, Sancho J, Bernabeu C. 1993. Structural and functional studies on the interaction of sodium dodecyl sulfate with ${\beta}$-galactosidase. Arch. Biochem. Biophys. 300: 451-457. https://doi.org/10.1006/abbi.1993.1061
  38. Muhsin TM, Hadi RB. 2001. Degradation of keratin substrates by fungi isolated from sewage sludge. Mycopathologia 154: 185-189.
  39. Patil CS, Gangawane AK, Hatti SS. 2010. Production and characterization of alkaline thermostable protease from newly isolated Bacillus sp. J. Plant Genom. 1: 9-17.
  40. Rai SK, Konwarh R, Mukherjee AK. 2009. Purification, characterization and biotechnological application of an alkaline b-keratinase produced by Bacillus subtilis RM-01 in solidstate fermentation using chicken-feather as substrate. Biochem. Eng. J. 45: 218-225. https://doi.org/10.1016/j.bej.2009.04.001
  41. Riffel A, Brandelli A. 2006. Keratinolytic bacteria isolated from feather waste. Braz. J. Microbiol. 37: 395-399. https://doi.org/10.1590/S1517-83822006000300036
  42. Riffel A, Lucas F, Heeb P, Brandelli A. 2003. Characterization of a new keratinolytic bacterium that completely degrades native feather keratin. Arch. Microbiol. 179: 258-265.
  43. Riffel A, Ortolan S, Brandelli A. 2003. De-hairing activity of extracellular proteases produced by keratinolytic bacteria. J. Chem. Technol. Biotechnol. 78: 855-859. https://doi.org/10.1002/jctb.828
  44. Saber WIA, El-Metwally MM, El-Hersh MS. 2010. Keratinase production and biodegradation of some keratinous wastes by Alternaria tenuissima and Aspergillus nidulans. Res. J. Microbiol. 5: 21-35. https://doi.org/10.3923/jm.2010.21.35
  45. Sangali S, Brandelli A. 2000. Feather keratin hydrolysis by Vibrio sp. strain kr2. J. Appl. Microbiol. 89: 735-743. https://doi.org/10.1046/j.1365-2672.2000.01173.x
  46. Sangali S, Brandelli A. 2000. Isolation and characterization of a novel feather-degrading bacterial strain. Appl. Biochem. Biotechnol. 87: 17-24. https://doi.org/10.1385/ABAB:87:1:17
  47. Santos RM, Firmino AA, de Sa CM, Felix CR. 1996. Keratinolytic activity of Aspergillus fumigatus Fresenius. Curr. Microbiol. 33: 364-370. https://doi.org/10.1007/s002849900129
  48. Singh J. 2002. Optimization of an extracellular protease of Chrysosporium keratinophilum and its potential in bioremediation of keratinic wastes. Mycopathologia 156: 151-156.
  49. Venugopal M, Saramma AV. 2006. Characterization of alkaline protease from Vibrio fluvialis strain VM 10 isolated from a mangrove sediment sample and its application as a laundry detergent additive. Process Biochem. 41: 1239-1243. https://doi.org/10.1016/j.procbio.2005.12.025
  50. 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. 222: 608-616.
  51. Wawrzkiewicz K, Wolski T, Lobarzewski J. 1991. Screening the keratinolytic activity of dermatophytes in vitro. Mycopathologia 114: 1-8. https://doi.org/10.1007/BF00436684
  52. Xie F, Chao Y, Yang X, Yang J, Xue Z, Luo Y, Qian S. 2009. Purification and characterization of four keratinases produced by Streptomyces sp. strain 16 in native human foot skin medium. Bioresour. Technol. 101: 344-350.

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