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A metallo-keratinase from a newly isolated Acinetobacter sp. R-1 with low collagenase activity and its biotechnological application potential in leather industry

Abstract

Microbial keratinase is a well-recognized enzyme that can specifically degrade insoluble keratins. A keratinase-producing bacterium was isolated from a duck ranch soil and identified as Acinetobacter sp. R-1 based on the biochemical characteristics and 16S rDNA gene sequencing. It showed high keratinase activity and low collagenase activity. The keratinase was purified to electrophoretic homogeneity with 6.69 % recovery, 2.68-fold purification and an estimated molecular weight of 25 kDa. Additionally, the keratinase showed optimal activity at 50 °C and pH11. Keratinase activity of Acinetobacter sp. significantly increased in the presence of Li+, Na+, and Ca2+, while it was completely inhibited by EDTA, indicating it was a metallo-keratinase. Moreover, the crude keratinase from Acinetobacter sp. R-1 could thoroughly depilate goat skin and simultaneously modify the wool surface, which indicated its applicable potential in leather and textile industries.

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References

  1. 1.

    Jaouadi B, Abdelmalek B, Fodil D, Ferradji FZ, Rekik H, Zarai N, Bejar S (2010) Purification and characterization of a thermostable keratinolytic serine alkaline proteinase from Streptomyces sp. strain AB1 with high stability in organic solvents. Bioresour Technol 101(21):8361–8369

  2. 2.

    Wang JJ, Garlich JD, Shih JCH (2006) Beneficial effects of versazyme, a keratinase feed additive, on body weight, feed conversion, and breast yield of broiler chickens. J Appl Poultry Res 15(4):544–550

  3. 3.

    George N, Sondhi S, Soni SK, Gupta N (2014) Lime and sulphide-free dehairing of animal skin using collagenase-free alkaline protease from Vibrio metschnikovii NG155. Indian J Microbiol 54(2):139–142

  4. 4.

    Saran S, Mahajan RV, Kaushik R, Isar J, Saxena RK (2013) Enzyme mediated beam house operations of leather industry: a needed step towards greener technology. J Clean Prod 54:315–322

  5. 5.

    Arunachalam C, Saritha K (2009) Protease enzyme an eco-friendly alternative for leather industry. Indian J Sci Technol 2(12):29–32

  6. 6.

    Shen JS, Rushforth M, Cavaco-Paulo A, Guebitz G, Lenting H (2007) Development and industrialisation of enzymatic shrink-resist process based on modified proteases for wool machine washability. Enzyme Microb Technol 40(7):1656–1661

  7. 7.

    Wang P, Wang Q, Fan X, Cui L, Yuan J, Chen S, Wu J (2009) Effects of cutinase on the enzymatic shrink-resist finishing of wool fabrics. Enzyme Microb Technol 44(5):302–308

  8. 8.

    Cardamone JM, Yao J, N A (2004) DCCA shrinkproofing of wool. Text Res J 74(6):550–560

  9. 9.

    Friedrich HGSKJ (2000) Keratinase of Doratomyces microsporus. Appl Microbiol Biotehnol 53:196–200

  10. 10.

    Tork SE, Shahein YE, El-Hakim AE, Abdel-Aty AM, Aly MM (2013) Production and characterization of thermostable metallo-keratinase from newly isolated Bacillus subtilis NRC 3. Int J Biol Macromol 55:169–175

  11. 11.

    Jaouadi NZ, Rekik H, Badis A, Trabelsi S, Belhoul M, Yahiaoui AB, Ben Aicha H, Toumi A, Bejar S, Jaouadi B (2013) Biochemical and molecular characterization of a serine keratinase from Brevibacillus brevis US575 with promising keratin-biodegradation and hide-dehairing activities. PLoS One 8(10):1–17

  12. 12.

    Bernal C, Cairó J, Coello N (2006) Purification and characterization of a novel exocellular keratinase from Kocuria rosea. Enzyme Microb Technol 38(1):49–54

  13. 13.

    Fang Z, Zhang J, Liu B, Du G, Chen J (2013) Biochemical characterization of three keratinolytic enzymes from Stenotrophomonas maltophilia BBE11-1 for biodegrading keratin wastes. Int Biodeter Biodegr 82:166–172

  14. 14.

    Jeong J-H, Lee OM, Jeon Y-D, Kim J-D, Lee N-R, Lee C-Y, Son H-J (2010) Production of keratinolytic enzyme by a newly isolated feather-degrading Stenotrophomonas maltophilia that produces plant growth-promoting activity. Process Biochem 45(10):1738–1745

  15. 15.

    Wu XQ, Chen L, Cao ZJ, Zhou MH (2012) Feather degradation and keratinase production by Stenotrophomonas maltophilia DHHJ. Adv Mater Res 550–553:1400–1403

  16. 16.

    Syed DG, Lee JC, Li WJ, Kim CJ, Agasar D (2009) Production, characterization and application of keratinase from Streptomyces gulbargensis. Bioresour Technol 100(5):1868–1871

  17. 17.

    Tatineni R, Doddapaneni KK, Potumarthi RC, Vellanki RN, Kandathil MT, Kolli N, Mangamoori LN (2008) Purification and characterization of an alkaline keratinase from Streptomyces sp. Bioresour Technol 99(6):1596–1602

  18. 18.

    Anitha TS, Palanivelu P (2013) Purification and characterization of an extracellular keratinolytic protease from a new isolate of Aspergillus parasiticus. Protein Expr Purif 88(2):214–220

  19. 19.

    Sivasubramanian S, Manohar BM, Puvanakrishnan R (2008) Mechanism of enzymatic dehairing of skins using a bacterial alkaline protease. Chemosphere 70(6):1025–1034

  20. 20.

    Shrinivas D, Naik GR (2011) Characterization of alkaline thermostable keratinolytic protease from thermoalkalophilic Bacillus halodurans JB 99 exhibiting dehairing activity. Int Biodeter Biodegr 65(1):29–35

  21. 21.

    Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227(5259):680–685

  22. 22.

    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(1–2):248–254

  23. 23.

    Yamamura S, Morita Y, Hasan Q, Rao SR, Murakami Y, Yokoyama K, Tamiya E (2002) Characterization of a new keratin-degrading bacterium isolated from deer fur. J Biosci Bioeng 93(6):595–600

  24. 24.

    Liu B, Zhang J, Li B, Liao X, Du G, Chen J (2013) Expression and characterization of extreme alkaline, oxidation-resistant keratinase from Bacillus licheniformis in recombinant Bacillus subtilis WB600 expression system and its application in wool fiber processing. World J Microbiol Biotechnol 29(5):825–832

  25. 25.

    Paul T, Halder SK, Das A, Bera S, Maity C, Mandal A, Das PS, Mohapatra PKD, Pati BR, Mondal KC (2013) Exploitation of chicken feather waste as a plant growth promoting agent using keratinase producing novel isolate Paenibacillus woosongensis TKB2. Biocatal Agr Biotechnol 2(1):50–57

  26. 26.

    Riffel A, Brandelli A (2002) Isolation and characterization of a feather-degrading bacterium from the poultry processing industry. J Ind Microbiol Biotehnol 29(5):255–258

  27. 27.

    Rajput R, Sharma R, Gupta R (2011) Cloning and characterization of a thermostable detergent-compatible recombinant keratinase from Bacillus pumilus KS12. Biotechnol Appl Bioc 58(2):109–118

  28. 28.

    Deng A, Wu J, Zhang Y, Zhang G, Wen T (2010) Purification and characterization of a surfactant-stable high-alkaline protease from Bacillus sp. B001. Bioresour Technol 101(18):7100–7106

  29. 29.

    Adel-El-Haleem D (2003) Acinetobacter environmental and biotechnological applications. Afr J Biotechnol 2(4):71–74

  30. 30.

    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(7):2325–2332

  31. 31.

    Chen K, Huang J, Chung C, Kuo W (2011) Identification and characterization of H10 enzymes isolated from Bacillus cereus H10 with keratinolytic and proteolytic activities. World J Microbiol Biotechnol 27(2):349–358

  32. 32.

    Gupta R, Ramnani P (2006) Microbial keratinases and their prospective applications: an overview. Appl Microbiol Biotechnol 70(1):21–33

  33. 33.

    Rai SK, Konwarh R, Mukherjee AK (2009) Purification, characterization and biotechnological application of an alkaline β-keratinase produced by Bacillus subtilis RM-01 in solid-state fermentation using chicken-feather as substrate. Biochem Eng J 45(3):218–225

  34. 34.

    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(3):169–173

  35. 35.

    Paul T, Das A, Mandal A, Halder SK, DasMohapatra PK, Pati BR, Mondal KC (2013) Biochemical and structural characterization of a detergent stable alkaline serine keratinase from Paenibacillus Woosongensis TKB2: a potential additive for laundry detergent. Waste Biomass Valori 5(4):563–574

  36. 36.

    Chaturvedi V, Bhange K, Bhatt R, Verma P (2014) Production of kertinases using chicken feathers as substrate by a novel multifunctional strain of Pseudomonas stutzeri and its dehairing application. Biocatal Agr Biotechnol 3(2):167–174

  37. 37.

    Pillai P, Archana G (2008) Hide depilation and feather disintegration studies with keratinolytic serine protease from a novel Bacillus subtilis isolate. Appl Microbiol Biotechnol 78(4):643–650

  38. 38.

    Jaouadi B, Ellouz-Chaabouni S, Ali MB, Messaoud EB, Naili B, Dhouib A, Bejar S (2009) Excellent laundry detergent compatibility and high dehairing ability of the Bacillus pumilus CBS alkaline proteinase (SAPB). Biotechnol Bioproc Eng 14(4):503–512

  39. 39.

    Dayanandan A, Kanagaraj J, Sounderraj L, Govindaraju R, Rajkumar GS (2003) Application of an alkaline protease in leather processing: an ecofriendly approach. J Clean Prod 11(5):533–536

  40. 40.

    Sundararajan S, Kannan CN, Chittibabu S (2011) Alkaline protease from Bacillus cereus VITSN04: potential application as a dehairing agent. J Biosci Bioeng 111(2):128–133

  41. 41.

    Jaouadi B, Ellouz-Chaabouni S, Rhimi M, Bejar S (2008) Biochemical and molecular characterization of a detergent-stable serine alkaline protease from Bacillus pumilus CBS with high catalytic efficiency. Biochimie 90(9):1291–1305

  42. 42.

    Rajkumar R, Jayappriyan KR, Rengasamy R (2011) Purification and characterization of a protease produced by Bacillus megaterium RRM2: application in detergent and dehairing industries. J Basic Microbiol 51(6):614–624

  43. 43.

    Paul T, Das A, Mandal A, Jana A, Maity C (2014) Effective dehairing properties of keratinase from Paenibacillus woosongensis TKB2 obtained under solid state fermentation. Waste Biomass Valor 5(1):97–107

  44. 44.

    Ismail A-MS, Housseiny MM, Abo-Elmagd HI, El-Sayed NH, Habib M (2012) Novel keratinase from Trichoderma harzianum MH-20 exhibiting remarkable dehairing capabilities. Int Biodeter Biodegr 70:14–19

  45. 45.

    Wang P, Wang Q, Cui L, Gao M, Fan X (2011) The combined use of cutinase, keratinase and protease treatments for wool bio-antifelting. Fiber Polym 12(6):760–764

  46. 46.

    Levene R, Cohen Y, Barkai D (1996) Applying proteases to confer improved shrink resistance to wool. J Soc Dyers Colour 112(1):6–10

  47. 47.

    Zhou D, He J (2006) Review of wool modification by protease. Dye Technol 28(8):12–14

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Acknowledgments

This work was financially supported by the National High Technology Research and Development Program of the People’s Republic China (No. 2012AA022204C), and the Ministry of Education of the People’s Republic of China (No. JUSRP51516).

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Correspondence to Jin-Song Shi or Zheng-Hong Xu.

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Zhang, R., Gong, J., Zhang, D. et al. A metallo-keratinase from a newly isolated Acinetobacter sp. R-1 with low collagenase activity and its biotechnological application potential in leather industry. Bioprocess Biosyst Eng 39, 193–204 (2016). https://doi.org/10.1007/s00449-015-1503-7

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Keywords

  • Acinetobacter sp. R-1
  • Keratinase
  • Purification
  • Dehairing
  • Surface modification