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Identification of Protease-Producing Halophilic Bacteria Isolated from Salt-Pond Soil
Corresponding Author(s) : Asep Awaludin Prihanto
Jurnal Ilmiah Perikanan dan Kelautan,
Vol. 12 No. 1 (2020): JURNAL ILMIAH PERIKANAN DAN KELAUTAN
Abstract
Highlights
- The halophilic bacteria were isolated from salt-pond soil.
- Isolate C2 was identified as the best protease producer.
- Isolate C2 was identified as B. amyloliquefaciens strain UBC
Abstract
Protease is an important enzyme for various industries, such as pharmaceuticals, leather production, meat processing, protein hydrolyzate, food products, and waste processing industries. This study aimed to isolate and identify protease-producing bacteria isolated from salt-pond soil at the Tuban Regency, East Java, Indonesia. Bacterial communities in the soil samples were firstly isolated from the soil samples by culture-dependent technique on Luria Bertani agar enriched with 5% NaCl. Thereafter, bacterial colonies that grew on the media were purified and screened for their protease production using a skim-milk agar. The bacterial colony which produced protease was further identified using phenotypic (gram staining) and genotypic assays (the 16S rDNA sequence). The result showed that one isolate out of six (isolate C2) obtained from the soil sample was observed to produce a protease enzyme. Based on its 16S rDNA sequence, the isolate was identified as Bacillus amyloliquefaciens strain UBC. These results suggest that B. Amyloliquefaciens strain UBC is a salt-tolerant bacterium (halophilic bacteria) which has the potential to be further developed for protease-producing biological agents.
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- Anbu, P. (2016). Enhanced Production and Organic Solvent Stability of A Protease from Brevibacillus laterosporus strain PAP04. Brazilian Journal of Medical and Biological Research, 49(4): e5178.
- Asokan, S., & Jayanthi, C. (2010). Alkaline Protease Production by Bacillus licheniformis and Bacillus coagulans. Journal of Cell & Tissue Research, 10(1): 2119.
- Choi, Y. H., Lee, J. S., Bae, S. Y., Yang, K. J., Yeom, K. W., Jo, D. H., Kang, O. H. & Baik, H. S. (2013). Isolation of Bacteria with Protease Activity from Cheonggukjang and Purification of Fibrinolytic Enzyme. Journal Life Science, 23(2):259-266.
- Choudhary, V., & Jain, P. C. (2012). Screening of Alkaline Protease Production by Fungal Isolates from Different Habitats of Sagar and Jabalpur District (M.P). Journal of Academia and Industrial Research, 1(4): 215-220.
- Dereeper, A., Guignon, V., Blanc, G., Audic, S., Buffet, S., Chevenet, F., Dufayard, J. F., Guindon, S., Lefort, V., Lescot, M., Claverie, J. M., & Gascuel, O. (2008). Phylogeny.fr: robust phylogenetic analysis for the non-specialist. Nucleic acids research, 36(Web Server issue), W465–W469. https://doi.org/10.1093/nar/gkn180
- Ebrahimpour, A., & Ashraf, K. (2015). Isolation, Characterization and Molecular Identification of Protease Producing Bacteria from Tashkooh Mountain Located in Ahvaz, Iran. International Journal of Life Sciences, 9 (2): 39–42.
- Gani, M. A., Tallei, T. E., & Fatimawali. 2019. Identifikasi Bakteri Asam Laktat dari Hasil Fermentasi Selada Romain (Lactuca sativa var. longifolia Lam.) Menggunakan Gen 16S rRNA. Jurnal Ilmiah Farmasi, 8(1): 57-65.
- Janda, J. M., & Abbott, S. L. (2007). 16S rRNA Gene Sequencing for Bacterial Identification in The Diagnostic Laboratory: Pluses, Perils, and Pitfalls. Journal of Clinical Microbiology, 45(9):2761-2764.
- Kim, M., Morrison, M., & Yu, Z. (2011). Evaluation of Different Partial 16S rRNA Gene Sequence Regions for Phylogenetic Analysis of Microbiomes. Journal of Microbiological Methods, 84(1), 81–87.
- Kumaunang, M., Sanchart, C., Suyotha, W., & Maneerat, S. (2019). Virgibacillus halodenitrificans MSK-10P, a Potential Protease-producing Starter Culture for Fermented Shrimp Paste (kapi) Production. Journal of Aquatic Food Product Technology. 28. 1-14. 10.1080/10498850.2019.1652874.
- Lane, D. J. (1991). 16S/23S rRNA sequencing, p. 115-175. In E. Stackebrandt and M. Goodfellow (ed.), Nucleic acid techniques in bacterial systematics. John Wiley & Sons, New York.
- Ma, S. C., Zhang, H. B., Ma, S. T., Wang, R., Wang, G. X., Shao, Y., & Li, C. X. (2015). Effects of Mine Wastewater Irrigation on Activities of Soil Enzymes and Physiological Properties, Heavy Metal Uptake and Grain Yield in Winter Wheat. Ecotoxicology and Environmental Safety, 113:483–490.
- Naiola, E., & Widhyastuti, N., 2002. Isolation, Selection and Optimalization of Protease Production of Some Bacterial Isolates. Berita Biologi, 6(3): 467-473.
- Nascimento, W. C. A. D., & Martins, M. L. L. (2006). Studies on The Stability Of Protease From Bacillus Sp. and Its Compatibility with Commercial Detergent. Brazilian Journal of Microbiology, 37(3): 307-311.
- Nursyam, H., & Prihanto, A. A. (2018). Identifikasi Molekuler Bakteri Endofit Mangrove Rhizopora Mucronata Penghasil Gelatinase (MMP2). Jurnal Pengolahan Hasil Perikanan Indonesia, 21(1):143-147.
- Prihanto, A. A., Aalifatul, F., Hartati, K., & Ken A. P. (2019). Identifikasi Bakteri Endofit Mangrove Api-Api Putih (Avicennia marina) Penghasil Enzim L-asparaginase. Jurnal Ilmiah Perikanan dan Kelautan, 10(2): 84-90.
- Sandhya, C., Sumantha, A., Szakacs, G., & Ashok P. (2005). Comparative Evaluation of Neutral Protease Production by Aspergillus oryzae in Submerged and Solid-State Fermentation. Process Biochemical, 40: 2689–2694.
- Sarker, P. K., Talukdar, S. A., Deb, P., Sayem, S. A., & Mohsina, K. (2013). Optimization and Partial Characterization of Culture Conditions for The Production of Alkaline Protease from Bacillus licheniformis P003. Springerplus, 2: 506.
- Singh, J. S. (2015). Microbes: The Chief Ecological Engineers in Reinstating Equilibrium in Degraded Ecosystems. Agriculture, Ecosystems & Environment, 203:80–82.
- Sinha, P., Singh, R.K., Srivastva, R., Sharma, R., & Tiwari, P. (2013). Characterization and Optimization of Alkaline Protease Enzyme Produced by Soil Borne Bacteria. Trends Life Science, 2(2): 2319–4731.
- Sun, J., Zhang, Q., Zhou, J., & Wei, Q. P. (2014). Illumina Amplicon Sequencing of 16S rRNA Tag Reveals Bacterial Community Development in The Rhizosphere of Apple Nurseries at A Replant Disease Site and A New Planting Site. Plos One, 9(10), e111744.
- Vishwanatha, K. S., Rao, A. A., & Singh, S. A. (2010). Acid Protease Production by Solid-State Fermentation Using Aspergillus oryzae MTCC 5341: Optimization of Process Parameters. Journal of Industrial Microbiology & Biotechnology, 37(2): 129-138.
- Wang, H., Yang, L., Ping, Y., Bai, Y., Luo, H., Huang, H., & Yao, B. (2016). Engineering of A Bacillus amyloliquefaciens Strain with High Neutral Protease Producing Capacity and Optimization of Its Fermentation Conditions. Plos One, 11(1), e0146373.
References
Anbu, P. (2016). Enhanced Production and Organic Solvent Stability of A Protease from Brevibacillus laterosporus strain PAP04. Brazilian Journal of Medical and Biological Research, 49(4): e5178.
Asokan, S., & Jayanthi, C. (2010). Alkaline Protease Production by Bacillus licheniformis and Bacillus coagulans. Journal of Cell & Tissue Research, 10(1): 2119.
Choi, Y. H., Lee, J. S., Bae, S. Y., Yang, K. J., Yeom, K. W., Jo, D. H., Kang, O. H. & Baik, H. S. (2013). Isolation of Bacteria with Protease Activity from Cheonggukjang and Purification of Fibrinolytic Enzyme. Journal Life Science, 23(2):259-266.
Choudhary, V., & Jain, P. C. (2012). Screening of Alkaline Protease Production by Fungal Isolates from Different Habitats of Sagar and Jabalpur District (M.P). Journal of Academia and Industrial Research, 1(4): 215-220.
Dereeper, A., Guignon, V., Blanc, G., Audic, S., Buffet, S., Chevenet, F., Dufayard, J. F., Guindon, S., Lefort, V., Lescot, M., Claverie, J. M., & Gascuel, O. (2008). Phylogeny.fr: robust phylogenetic analysis for the non-specialist. Nucleic acids research, 36(Web Server issue), W465–W469. https://doi.org/10.1093/nar/gkn180
Ebrahimpour, A., & Ashraf, K. (2015). Isolation, Characterization and Molecular Identification of Protease Producing Bacteria from Tashkooh Mountain Located in Ahvaz, Iran. International Journal of Life Sciences, 9 (2): 39–42.
Gani, M. A., Tallei, T. E., & Fatimawali. 2019. Identifikasi Bakteri Asam Laktat dari Hasil Fermentasi Selada Romain (Lactuca sativa var. longifolia Lam.) Menggunakan Gen 16S rRNA. Jurnal Ilmiah Farmasi, 8(1): 57-65.
Janda, J. M., & Abbott, S. L. (2007). 16S rRNA Gene Sequencing for Bacterial Identification in The Diagnostic Laboratory: Pluses, Perils, and Pitfalls. Journal of Clinical Microbiology, 45(9):2761-2764.
Kim, M., Morrison, M., & Yu, Z. (2011). Evaluation of Different Partial 16S rRNA Gene Sequence Regions for Phylogenetic Analysis of Microbiomes. Journal of Microbiological Methods, 84(1), 81–87.
Kumaunang, M., Sanchart, C., Suyotha, W., & Maneerat, S. (2019). Virgibacillus halodenitrificans MSK-10P, a Potential Protease-producing Starter Culture for Fermented Shrimp Paste (kapi) Production. Journal of Aquatic Food Product Technology. 28. 1-14. 10.1080/10498850.2019.1652874.
Lane, D. J. (1991). 16S/23S rRNA sequencing, p. 115-175. In E. Stackebrandt and M. Goodfellow (ed.), Nucleic acid techniques in bacterial systematics. John Wiley & Sons, New York.
Ma, S. C., Zhang, H. B., Ma, S. T., Wang, R., Wang, G. X., Shao, Y., & Li, C. X. (2015). Effects of Mine Wastewater Irrigation on Activities of Soil Enzymes and Physiological Properties, Heavy Metal Uptake and Grain Yield in Winter Wheat. Ecotoxicology and Environmental Safety, 113:483–490.
Naiola, E., & Widhyastuti, N., 2002. Isolation, Selection and Optimalization of Protease Production of Some Bacterial Isolates. Berita Biologi, 6(3): 467-473.
Nascimento, W. C. A. D., & Martins, M. L. L. (2006). Studies on The Stability Of Protease From Bacillus Sp. and Its Compatibility with Commercial Detergent. Brazilian Journal of Microbiology, 37(3): 307-311.
Nursyam, H., & Prihanto, A. A. (2018). Identifikasi Molekuler Bakteri Endofit Mangrove Rhizopora Mucronata Penghasil Gelatinase (MMP2). Jurnal Pengolahan Hasil Perikanan Indonesia, 21(1):143-147.
Prihanto, A. A., Aalifatul, F., Hartati, K., & Ken A. P. (2019). Identifikasi Bakteri Endofit Mangrove Api-Api Putih (Avicennia marina) Penghasil Enzim L-asparaginase. Jurnal Ilmiah Perikanan dan Kelautan, 10(2): 84-90.
Sandhya, C., Sumantha, A., Szakacs, G., & Ashok P. (2005). Comparative Evaluation of Neutral Protease Production by Aspergillus oryzae in Submerged and Solid-State Fermentation. Process Biochemical, 40: 2689–2694.
Sarker, P. K., Talukdar, S. A., Deb, P., Sayem, S. A., & Mohsina, K. (2013). Optimization and Partial Characterization of Culture Conditions for The Production of Alkaline Protease from Bacillus licheniformis P003. Springerplus, 2: 506.
Singh, J. S. (2015). Microbes: The Chief Ecological Engineers in Reinstating Equilibrium in Degraded Ecosystems. Agriculture, Ecosystems & Environment, 203:80–82.
Sinha, P., Singh, R.K., Srivastva, R., Sharma, R., & Tiwari, P. (2013). Characterization and Optimization of Alkaline Protease Enzyme Produced by Soil Borne Bacteria. Trends Life Science, 2(2): 2319–4731.
Sun, J., Zhang, Q., Zhou, J., & Wei, Q. P. (2014). Illumina Amplicon Sequencing of 16S rRNA Tag Reveals Bacterial Community Development in The Rhizosphere of Apple Nurseries at A Replant Disease Site and A New Planting Site. Plos One, 9(10), e111744.
Vishwanatha, K. S., Rao, A. A., & Singh, S. A. (2010). Acid Protease Production by Solid-State Fermentation Using Aspergillus oryzae MTCC 5341: Optimization of Process Parameters. Journal of Industrial Microbiology & Biotechnology, 37(2): 129-138.
Wang, H., Yang, L., Ping, Y., Bai, Y., Luo, H., Huang, H., & Yao, B. (2016). Engineering of A Bacillus amyloliquefaciens Strain with High Neutral Protease Producing Capacity and Optimization of Its Fermentation Conditions. Plos One, 11(1), e0146373.