Int. J. Biosci.11(4), 228-236, October 2017
Extracellular proteases have a fundamental position with respect to their physiological roles as well as their commercial applications. Bacillus licheniformis RT7P1, was evaluated in this study for production of extracellular protease activity. The culture was maintained on 1% w/v casein plates. In this study, the strain was found to produce maximum enzyme at pH 7 and temperature 37°C after 72 h. The optimum assay pH and the temperature was 10 and 50°C, respectively. The enzyme was stable between pH 9-11and thermal stability data showed that enzyme was stable at 100°C. For cloning of protease gene from Bacillus licheniformis, primers were designed to pick their full-length sequences from the genomic DNA obtained from different Bacillus species. Genomic DNA was isolated from Bacillus licheniformis strain RT7P1 and then the protease gene was amplified from it by using RT7P1 specific primers. This amplified product (1725bp) was then cloned in PTZ57R/T vector. The clone was confirmed by restriction analysis with EcoR1 and BamH1, which showed two fragments of (2886 bp) and (1725 bp), which showed that insert was cloned in the right orientation. The study indicates that Bacillus licheniformis RT7P1 is a good source of commercial thermostable alkaline extracellular protease.
Singhal P, Nigam VK, Vidyarthi AS. 2012. Studies on production, characterization, and application of microbial alkaline proteases. Nature, 3, 653-669.
Boominadhan U, Rajakumar R, Karpaga P, Sivakumaar V, Joe MM. 2009. Optimization of protease enzyme production using Bacillus Sp. isolated from different wastes. International Journal of Botany and Research 2, 83-87 p.
Morya V, Yadav D. 2009. Production and partial characterization of neutral protease by an indigenously isolated strain of Aspergillus tubingensis NIICC-08155. International Journal of Microbiology 8, 1-6.
Lagzian M, Asoodeh A. 2012. An extremely thermotolerant, alkaliphilic subtilisin-like protease from hyperthermophilic Bacillus sp. MLA64. International Journal of Biological Macromolecules 51, 960–967 p.
Cotarleţ M, Negoita T, Bahrim G, Stougaard P. 2009. Cold adapted amylase and protease from new streptomyces 4 alga antarctic strain. Journal of Innovative Food Science and Emerging Technologies, 5, 23-30 p
Suganthi C, Mageswari A, Karthikeyan, S, Anbaagan M, Sivakumar A, Gothandam KM. 2013. Screening and optimization of protease production from a halotolerant Bacillus licheniformis isolated from saltern sediments. Journal of Genetic Engineering and Biotechnology 11, p 47-52.
Pant G, Prakash O, Chandra M, Sethi S, Punetha1 H, Dixit S, Pant AK. 2014. Biochemical analysis, pharmacological activity, antifungal activity and mineral analysis in methanolic extracts of Myrica esculenta and Syzygium cumini: the Indian traditional fruits growing in Uttarakh and Himalaya. Indian Journal of Pharmaceutical and Biological Research, 2(1), 26-34 p.
Mathew CD, Gunathilaka RMS. 2015. Production purification and characterization of thermostable alkaline serine protease from Bacillus licheniformis NMS-1. International Journal of Biotechnology and Molecular Biology Research, 6(3), 19-27 p.
Wilson P, Remigio Z. 2012. Production and characterization of protease enzyme produced by a novel moderate thermophilic bacterium (EP1001) isolated from an alkaline hot spring, Zimbabwe. African Journal of Microbiology Research, 6(27), 5542-5551 p.
Talebi M, Emtiazi G, Sepahy AA, Zaghian, S. 2013. Zymogram analysis of alkaline keratinase produced by nitrogen fixing Bacillus pumilus ZED17 exhibiting multiprotease enzyme activities. The journal of Microbiology 6(10), 1-6 p.
Gärtner D, Geissendörfer M, Hillen W. 1988. Expression of Bacillus subtilis xyl operon is repressed at the level of transcription and is induced by xylose. Journal of Bacteriology, 170 (7), 3102–3109 p.
Sayem SMA, Alam MJ, Hoq MM. 2006. Effect of temperature, pH and metal ions on the activity and stability of alkaline protease from novel Bacillus licheniformis MZK03. Proceedings of the Pakistan Academy of Sciences 43(4), 257-262 p.
Bernlohr RW, Clark V. 1971. Characterization and regulation of protease synthesis and sctivity in Bacillus licheniformis. Journal of Bacteriology, vol 105(1), 276–283 p.
Joo HS, Choi JW. 2012. Purification and characterization of a novel alkaline protease from Bacillus horikoshii”, Journal of Microbiology and Biotechnology 22 (1), 58-68 p.
Rozs M, Manczinger L, Vagvolgyi LC, Kevei, 2001. Secretion of a trypsin-like thiol protease by a new keratinolytic strain of Bacillus licheniformis. FEMS Microbiology Letter, 205(2), 221-4 p.
Ferrero MA, Castro GR, Abate CM, Bagori MD, Sinerizf. 1996. Isolation, production and characterization of thermostable alkaline protease of Bacillus licheniformis MIR29. Journal of Applied Microbialogy and Biotechnology 45, 327-332 p.
Guangrong H, Tiejing Y, Jiaxing HPO. 2006. Purification and characterization of a protease from thermophilic Bacillus strain HS08. African Journal of Biotechnology 5, 2433-2438 p.
Nascimento CAD, Martin MLL. 2006. Studies on the stability of protease from Bacillus sp. and its compatibility with commercial detergents. Brazillian Journal of Microbiology37, 307-311 p.
Adinarayana K, Ellaiah P, Prasad DS. 2003. Purification and partial characterization of thermostable serine alkaline protease from a newly isolated Bacillus subtilis PE-11”, AAPS. Pharmaceutical Science and Technology 4, 440-448 p.
Johnston J, Pippen X, Pivot M., Lichinitser S, Sadeghi V, Dieras HL, Gomez G, Romieu A, Manikhas, Kennedy MJ. 2009. Lapatinib combined with letrozole versus letrozole and placebo as first line therapy for postmenopausal hormone receptor positive metastatic breast cancer. Journal of Clinical Oncology 27, 5538-5546 p.
Rasool N, Rashid N, Javed MA, Haider MS. 2011. Requirement of pro-peptide in proper folding of Subtilisin-like serine protease TK0076. Pakistan Journal of Botany 43, 2059-2065.