Biotechnology and Bioprocess Engineering 2020; 25(1): 104-116  
Endoglucanase Produced by Bacillus subtilis Strain CBS31: Biochemical Characterization, Thermodynamic Study, Enzymatic Hydrolysis, and Bio-industrial Applications
Sudip Regmi1,†, Yoon Seok Choi1,†, Young Kyun Kim1,†, Md Maruf Khan1, Sang Hun Lee1, Seung Sik Cho2, Ying-Yu Jin3, Dae Young Lee4, Jin Cheol Yoo1,*, and Joo-Won Suh3,*
1Department of Pharmacy, College of Pharmacy, Chosun University, Gwangju 61452, Korea
2Department of Pharmacy, Mokpo National University, Muan 58554, Korea
3Center for Nutraceutical and Pharmaceutical Materials, Myongji University, Yongin 17058, Korea
4Department of Herbal Crop Research, National Institute of Horticultural and Herbal Science, RDA, Eumseong 27709, Korea
Correspondence to: Jin Cheol Yoo*
Department of Pharmacy, College of Pharmacy, Chosun University, Gwangju 61452, Korea
Tel: +82-62-230-6380; Fax: +82-62-227-3963
E-mail: jcyu@chosun.ac.kr
Joo-Won Suh*
Center for Nutraceutical and Pharmaceutical Materials, Myongji University, Yongin 17058, Korea
Tel: +82-31-330-6190; Fax: +82-31-336-0870
E-mail: jwsuh@mju.ac.kr

Authors contributed equally to this work.
Received: August 26, 2019; Revised: October 13, 2019; Accepted: October 16, 2019; Published online: February 29, 2020.
© The Korean Society for Biotechnology and Bioengineering. All rights reserved.

This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/3.0/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.
Abstract
Microbial cellulases have become the mainstream biocatalysts due to their complex nature and widespread industrial applications. Here, homogeneous endoglucanase GluCB31 from Bacillus subtilis subsp. inaquosorum CBS31 was studied. GluCB31 was purified to 17.68-fold with an 8.33% yield and a specific activity of 1066.37 U/mg. Biochemical properties of GluCB31 were performed and the results are as follows; molecular mass of 35 kDa with an optimum pH at 7.5 and temperature at 50oC. GluCB31 was immobilized in calcium alginate gel and it exhibited the highest activity at 10oC higher temperature than soluble enzyme, as the entrapment in alginate gel made GluCB31 more stable. Kinetic studies showed the Vmax of 1293.33 ± 2.51 U/mg and Km of 0.0183 mg/mL. Enzymatic activity was activated by Tween-20 (106.7%), Tween-80 (111.6%), Triton X-100 (142.3%), SDS (135.5%), Mg++ (185.7%), Cu++ (167.6%), Zn++ (153.7%), Mn++ (106.3%), Ba++ (181.9%), Ni++ (107.2%) while inhibited by Fe++ (15.8%), β-mercaptoethanol (46.8%), EDTA (54.5%). Enthalpy, free energy, and entropy of activation were calculated to be 38.526 kJmol-1, 44.187 kJmol-1, and -17.518 Jmol-1K-1 respectively. Also, ΔGE-S and ΔGE-T were found to be -10.75 kJmol-1 and -45.92 kJmol-1 respectively. A low ΔS, ΔGE-S, and ΔGE-T values were signified enzyme-catalyzed reaction occurs at a fast rate and the existence of the enzyme in its stable state. Cellobiose was the major end product of hydrolysis. These attributes of GluCB31 demonstrated the diversity of catalytic activities and serve in various biotechnological processes, thus deserve to be developed as a bio-industrial agent.
Keywords: endoglucanase, thermodynamic, bio-industrial, enzymatic hydrolysis


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