Optimization conditions of alkaline protease production by Streptomyces sp.H1 isolated from red sea coastal region in submerged culture

  • Authors

    • Hind A. A. Al-Zahrani 1Biology Depart., Faculty of Sciences, University of Jeddah, Jeddah, Saudi Arabia
  • 16S Rdna, Alkaline Protease, Optimization, Streptomyces SP, Submerged Culture.
  • A potent alkaline protease producing strain characterized and identified as Streptomyces sp. H1 was isolated from soil around red sea shore. The enzyme was produced extracellulary in submerged culture revealed maximum level during early stationary phase. Alkaline protease showed the highest activity at incubation time, pH and inoculum size of 3 days, 9 and 8% respectively. Among different carbon sources beet molasses gave a maximum production followed by starch, sucrose and fructose. High yield of protease production was noticed with casein followed by peptone, yeast extract and ammonium sulphate. Furthermore, it was optimized with 7g/l NaCl resulted in higher level of protease. Optimization of the process parameters resulted in about 3.4 fold increase in the alkaline protease. Partial purification of the crude enzyme was achieved by fractional precipitation using ammonium sulfate at 50% saturation. Due to the maximum production of protease in the presence of cheaper substrate as beet molasses, stability at alkaline pH 9 and temperature up to 70 oC besides salt tolerance make the strain and its enzyme useful in different industrial applications.



  • References

    1. [1] Suthindhiran K, Jayasri M, Dipali D and Prasar A (2014).Screening and characterization of protease producing actinomycetes from marine saltern. Journal of Basic Microbiology, 54(10): 1098-1109. https://doi.org/10.1002/jobm.201300563.

      [2] Singhal P, NigamV K, Vidyarthi AS (2012). Studies on production characterization and applications of microbial alkaline proteases. International Journal of Advanced Biotechnology Research, 3: 653- 669.

      [3] Mukesh Kumar DJ, Premavathi V, Govindarajan N, Balakumaran MD, Kalaichelvan PT (2012). Production and purification of alkaline protease from Bacillus sp. MPTK 712 isolated from dairy sludge. Global Veterinaria, 8: 433-439.

      [4] El-Shafei H, Abdel-Aziz M, Ghali M and Abdalla A. (2010). Proceeding of Fifth Scientific Environmental Conference, ZagazigUniv, 125-142.

      [5] Ramesh S, Rajesh M and Mathivanan N (2009). Characterization of a thermostable alkaline protease produced by marine Streptomyces fungicidicus MML1614. Bioprocess and Biosystems Engineering, 32: 791. https://doi.org/10.1007/s00449-009-0305-1.

      [6] Okpukpara O B and George-Okafor U O (2016) .Production of Thermostable Alkaline Protease from Streptomyces sp-A54. Nigerian Journal of Biotechnology, 31: 88 – 95. https://doi.org/10.4314/njb.v31i1.12.

      [7] Jayasree D, Sandhya Kumara TD, Kavi Kishor PB, Vi- jaya Lakshmi M, Lakshmi Narasu M. (2009). Optimization of production protocol of alkaline protease by Streptomyces pulvereceus. International JRI of Science and Technology, 1: 79- 82.

      [8] Arifuzzaman M, Khatun M R and Rahman H (2010). Isolation and screening of actinomycetes from sundarbans soil for antibacterial activity. African Journal of Biotechnology, 9:4615-4619.

      [9] Shirling EB, Gottlieb D (1966). Methods for characterization of Streptomyces species. International journal of systematic bacteriology, 16: 313-340. https://doi.org/10.1099/00207713-16-3-313.

      [10] Pridham TG (1964). Color of Streptomyces: report of an international workshop on determination of color of Streptomyces. Applied Microbiology, 13: 43-6.

      [11] Kawato M, Shinobu R (1959). Cover slip culture of Streptomyces herbaricolour nov. Sp. supplement; a simple technique for the microscopic observation. Mem. Osaka University, Lib. Art. Educ. 8: 114-119.

      [12] Bergey’s manual of determinative bacteriology (2000). Actinomycetales. 9th edition.

      [13] Sambrook J, Fritsch E F, Maniaties T (1989) Molecular Cloning: A Laboratory Manual second Cold. Spring, Harbor Laboratory press, Cold Spring Harbor, New York, USA.

      [14] Hall TA (1999). BioEdit: a user-friendly biological sequence alignment editor and analysis program for Windows 95/98/NT. Nucleic Acids Symposium Series, 41: 95-98.

      [15] Tsuchida O, Yamagota Y, Ishizuka J, Arai J, Yamada J, Takeuchi M, Ichishima E (1986) . An alkaline protease of an alkalophilic Bacillus sp. Current Microbiology, 14:7-12. https://doi.org/10.1007/BF01568094.

      [16] Lowry OH, Rosebrough NJ, Farr AL, Randall RJ. (1951). Protein measurement with Folin phenol reagent. Journal of Biological Chemistry, 193:265-275.

      [17] Bradford M M (1976). A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Analytical Biochemistry, 72:248-254. https://doi.org/10.1016/0003-2697(76)90527-3.

      [18] Strzelczyk E and Szpotański T (1989). Cellulolytic and pectolytic activity of streptomycetes isolated from root-free soil, rhizosphere and mycorrhizosphere of pine (Pinus sylvestris L.). Biology and Fertility of Soils, 9: 268-272. https://doi.org/10.1007/BF00336238.

      [19] Al-Askar, A. A., Rashad, Y. M., Hafez, E. E., Abdulkhair, W. M., Baka, Z. A., & Ghoneem, K. M. (2015a). Characterization of alkaline protease produced by Streptomyces griseorubens E44G and its possibility for controlling Rhizoctonia root rot disease of corn. Biotechnology & Biotechnological Equipment, 29(3), 457–462. https://doi.org/10.1080/13102818.2015.1015446.

      [20] Kitadokoro K, Tsuzuki H, Nakamura E , Sato T, Teraoka H (1994). Purification, characterization, primary structure, crystallization and preliminary crystallographic study of a serine proteinase from Streptomyces fradiae ATCC 14544, European Journal of Biochemistry, 220: (1) 55–61. https://doi.org/10.1111/j.1432-1033.1994.tb18598.x.

      [21] Korkare CR, Mahadik KR, Kadam SS, Chopade BA (2004) Isolation, characterization and antimicrobial activity of marine halophilic Actinopolyspora species AH1 from the West coast of India. Current Sciences, 86593–597.

      [22] Holt JG, Krieg NR, Sneath PH, et al. Group 19. Regular, non-sporing Gram-positive rods. In: Hensyl WR (ed), editor (1994). Bergey's manual of determinative bacteriology. Ninth ed. Baltimore: Williams & Wilkins Inc. 565–70.

      [23] Moreira K A, Cavalcanti M T, Duarte H S, Tambourgi E B, Silva VL, Porto AN L, Filho J L (2001). Partial characterization of proteases from Streptomyces clavuligerus using an inexpensive medium. Brazilian Journal of Microbiology, 32:23–28. https://doi.org/10.1590/S1517-83822001000300010.

      [24] El-Sayed E M, Moataza M S, Hassan M. A, Mohsen H S, and Helmy M. H (2012). Optimization Conditions of Extracellular Proteases Production from a Newly Isolated Streptomyces Pseudogrisiolus NRC-15. E-Journal of Chemistry, 9: (2) 949-961. https://doi.org/10.1155/2012/168540.

      [25] Haritha R, SivaKumar K, Swathi A, Jagan Mohan YSYV and Ramana T (2012). Characterization of marine Streptomyces carpaticus and optimization of conditions for production of extracellular protease. Microbiology Journal, 2: 23-35. https://doi.org/10.3923/mj.2012.23.35.

      [26] Chi Z M and Zhao S Z (2003). Optimization of medium and cultivation conditions for pullulan production by new pullulan-producing yeast strain. Enzyme and Microbial Technology, 33: 206-211. https://doi.org/10.1016/S0141-0229(03)00119-4.

      [27] Lotfy WA (2007). Production of cephalosporin C by Acremonium chrysogenum grown on beet molasses: Optimization of process parameters through statistical experimental designs. Research Journal of Microbiology, 2: 1-12. https://doi.org/10.3923/jm.2007.1.12.

      [28] De Azeredo LAI, De Lima MB, Coelho RRR and Freire DMG (2006). A lowâ€cost fermentation medium for thermophilic protease production by Streptomyces sp 594 using feather meal and corn steep liquor. Current Microbiology, 53: 335–339. https://doi.org/10.1007/s00284-006-0163-x.

      [29] Thumar JT, Singh SP (2007). Secretion of an alkaline protease from salt-tolerant and alkaliphilic, Streptomyces clavuligerus strain Mit-1. Brazilian Journal of Microbiology, 38:1–9 https://doi.org/10.1590/S1517-83822007000400033.

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    A. A. Al-Zahrani, H. (2018). Optimization conditions of alkaline protease production by Streptomyces sp.H1 isolated from red sea coastal region in submerged culture. International Journal of Basic and Applied Sciences, 7(3), 48-54. https://doi.org/10.14419/ijbas.v7i3.14519