A multi-tolerant low molecular weight mannanase from Bacillus sp. CSB39 and its compatibility as an industrial biocatalyst |
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| Institution: | 1. Department of Pharmacy, College of Pharmacy, Chosun University, Gwangju 501-759, Republic of Korea;2. Department of Pharmacy, Mokpo National University, Muan, Jeonnam 534-729, Republic of Korea;1. Department of Chemical and Process Engineering, Faculty of Engineering and Built Environment, Universiti Kebangsaan Malaysia, 43600 Bangi, Selangor, Malaysia;2. School of Bioprocess Engineering, Universiti Malaysia Perlis, Kompleks Pusat Pengajian Jejawi 3, 02600 Arau, Perlis, Malaysia;3. Department of Microbiology, Masjed Soleyman Branch, Islamic Azad University, Masjed Soleyman, Iran;4. Department of Applied Microbiology, Faculty of Applied Sciences, Taiz University, 6803 Taiz, Yemen;5. School of Biosciences and Biotechnology, Faculty of Sciences and Technology, Universiti Kebangsaan Malaysia, 43600 Bangi, Selangor, Malaysia;1. State Key Laboratory of Bioreactor Engineering and Institute of Applied Chemistry, East China University of Science and Technology, Shanghai 200237, PR China;2. Collaborative Innovation Center for Biomanufacturing Technology, Shanghai 200237, PR China;1. Department of Protein Chemistry and Technology, CSIR-Central Food Technological Research Institute, Mysuru 570 020, India;2. Academy of Scientific and Innovative Research (AcSIR), CSIR-CFTRI Campus, Mysuru 570 020, India;1. Department of Microbiology, Dr. Harisingh Gour Vishwavidyalaya (A Central University), Sagar, MP 470003, India;2. Department of Physics, Dr. Harisingh Gour Vishwavidyalaya (A Central University), Sagar, MP 470003, India;1. Department of Biochemistry and Microbiology, Rhodes University, Grahamstown 6140, Eastern Cape, South Africa;2. Forest Products Biotechnology, University of British Columbia, 2424 Main Mall, Vancouver, British Columbia, Canada V6T1Z4 |
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| Abstract: | Bacillus sp. CSB39, isolated from popular traditional Korean food (Kimchi), produced a low molecular weight, thermostable mannanase (MnCSB39); 571.14 U/mL using locust bean gum galactomannan as a major substrate. It was purified to homogeneity using a simple and effective two-step purification strategy, Sepharose CL-6B and DEAE Sepharose Fast Flow, which resulted in 25.47% yield and 19.32-fold purity. The surfactant-, NaCl-, urea-, and protease-tolerant monomeric protein had a mass of ∼30 kDa as analyzed by SDS-PAGE and galactomannan zymography. MnCSB39 was found to have optimal activity at pH 7.5 and temperature of 70 °C. The enzyme showed ˃55% activity at 5.0–15% (w/v) NaCl, and ˃93% of the initial activity after incubation at 37 °C for 60 min. Trypsin and proteinase K had no effect on MnCBS39. The enzyme showed ˃80% activity in up to 3 M urea. The N-terminal amino acid sequence, ALKGDGX, did not show identity with reported mannanases, which suggests the novelty of our enzyme. Activation energy for galactomannan hydrolysis was 26.85 kJmol−1 with a Kcat of 142.58 × 104 s−1. MnCSB39 had Km and Vmax values of 0.082 mg/mL and 1099 ± 1.0 Umg−1, respectively. Thermodynamic parameters such as ΔH, ΔG, ΔS, Q10, ΔGE-S, and ΔGE-T supported the spontaneous formation of products and the high hydrolytic efficiency and feasibility of the enzymatic reaction, which strengthen its novelty. MnCSB39 activity was affected by metal ions, modulators, chelators, and detergents. Mannobiose was the principal end-product of hydrolysis. Bacillus subtilis CSB39 produced a maximum of 1524.44 U mannanase from solid state fermentation of 1 g wheat bran. MnCSB39 was simple to purify, was active at a wide pH and temperature range, multi-stress tolerant and catalyzes a thermodynamically possible reaction, characteristics that suggests its suitability for application as an industrial biocatalyst. |
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| Keywords: | Biocatalyst Biodegradation Galactomannan Mannanase |
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