title: A multi-tolerant low molecular weight mannanase from Bacillus sp. CSB39 and its compatibility as an industrial biocatalyst.
abstract: Bacillus sp. CSB39, isolated from popular traditional Korean food (Kimchi), produced a low molecular weight, thermostable mannanase (MnCSB39); 571.14U/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 ∼30kDa 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 60min. Trypsin and proteinase K had no effect on MnCBS39. The enzyme showed ˃80% activity in up to 3M 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 26.85kJmol(-1) with a Kcat of 142.58×10(4)s(-1). MnCSB39 had Km and Vmax values of 0.082mg/mL and 1099±1.0Umg(-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 1524.44U mannanase from solid state fermentation of 1g 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.
authors.affiliation: Department of Pharmacy, College of Pharmacy, Chosun University, Gwangju 501-759, Republic of Korea.
authors.forename: Sudip
authors.initials: S
authors.lastname: Regmi
authors.affiliation: Department of Pharmacy, College of Pharmacy, Chosun University, Gwangju 501-759, Republic of Korea.
authors.forename: Pradeep
authors.initials: P
authors.lastname: G C
authors.affiliation: Department of Pharmacy, College of Pharmacy, Chosun University, Gwangju 501-759, Republic of Korea.
authors.forename: Yun Hee
authors.initials: YH
authors.lastname: Choi
authors.affiliation: Department of Pharmacy, College of Pharmacy, Chosun University, Gwangju 501-759, Republic of Korea.
authors.forename: Yoon Seok
authors.initials: YS
authors.lastname: Choi
authors.affiliation: Department of Pharmacy, College of Pharmacy, Chosun University, Gwangju 501-759, Republic of Korea.
authors.forename: Ji Eun
authors.initials: JE
authors.lastname: Choi
authors.affiliation: Department of Pharmacy, Mokpo National University, Muan, Jeonnam 534-729, Republic of Korea.
authors.forename: Seung Sik
authors.initials: SS
authors.lastname: Cho
authors.affiliation: Department of Pharmacy, College of Pharmacy, Chosun University, Gwangju 501-759, Republic of Korea. Electronic address: jcyu@chosun.ac.kr.
authors.forename: Jin Cheol
authors.initials: JC
authors.lastname: Yoo
mesh_terms: D001412:Bacillus subtilis / Q000201:enzymology* / Q000235:genetics / Q000302:isolation & purification; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D055162:Biocatalysis; D004795:Enzyme Stability; D005516:Food Microbiology; D005685:Galactans; D007218:Industrial Microbiology; D007700:Kinetics; D008351:Mannans; D008970:Molecular Weight; D053149:Plant Gums; D013379:Substrate Specificity; D013816:Thermodynamics; D044902:beta-Mannosidase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D005685:Galactans; D008351:Mannans; D053149:Plant Gums; D044902:beta-Mannosidase; C017471:locust bean gum
keywords: Bacillus sp.; Biocatalyst; Biodegradation; Galactomannan; Mannanase