Purification and identification of novel alkaline pectinase PNs31 from Bacillus subtilis CBS31 and its immobilization fo
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pISSN: 0256-1115 eISSN: 1975-7220
INVITED REVIEW PAPER
INVITED REVIEW PAPER
Purification and identification of novel alkaline pectinase PNs31 from Bacillus subtilis CBS31 and its immobilization for bioindustrial applications Md. Saifur Rahman*,‡, Young Kyun Kim*,‡, Md Maruf Khan*, Sang Hun Lee*, Yun Hee Choi*, Seung Sik Cho**, Chulhwan Park***,†, and Jin Cheol Yoo*,† *Department of Pharmacy, College of Pharmacy, Chosun University, 309 Pilmun-daero, Dong-gu, Gwangju 61452, Korea **Department of Pharmacy, Mokpo National University, 1666 Yeongsan-ro, Muan-gun, Jeollanam-do 58554, Korea ***Department of Chemical Engineering, Kwangwoon University, 20 Kwangwoon-ro, Nowon-gu, Seoul 01897, Korea (Received 4 June 2020 • Revised 23 July 2020 • Accepted 27 July 2020) AbstractA robust alkaline pectinase PNs31 was produced from Bacillus subtilis CBS31 that was isolated from kimchi (a traditional Korean fermented food). The isolated strain CBS31 was identified as having 99.93% similarity to Bacillus subtilis subsp. The PNs31 was purified to homogeneity using two-step column purification, with an estimated 13.6% pectinase yield and 29.2-fold purity. The molecular mass (~35 kDa) and the N-terminal sequence residues of PNs31 established it as a novel pectinolytic enzyme. PNs31 was stable in the alkaline pH range (9-14) and up to 70 oC and had an optimum pH and temperature were 12.2 and 60 oC, respectively. PNs31 had definite Vmax and Km values of 220.43 mmol/min and 0.42 mg/mL, respectively, when different concentrations (5 to 50 mg/mL) of pectin were used as substrate. To develop the functional stability and reusability of the pectinase enzyme, PNs31 was immobilized in calcium alginate beads. The result was that the immobilized enzyme maintained approximately 83% and 65% relative activity in the second and third cycles of reusability experiments, respectively. Keywords: Alkaline Pectinase, Bacillus subtilis, Biochemical Characterization, Immobilization, Purification
eration. In particular, fermented products can be used directly in the process as crude enzymes without purification. However, commercial bulk proteins are mass-produced through submerged culture system as scale-up is difficult and requires a lot of labor [7]. Despite the superior catalytic activity of pectinase, the use of free enzymes constantly presents the following problems for industrial applications: additional product recovery steps, low stability of the enzyme, and limited reusability [8]. These can be overcome through enzyme immobilization, which can improve catalyst stability and help in continuous reuse of expensive catalysts. We can use bound protein for further process with repeated use as well as it can be released by treatment. Thus, industrial application of the produced enzyme can be enhanced [9-11]. In this study, an active pectinase (PNs31) was purified from a Bacillus strain, and its biochemical characteristics were investigated. In addition, the immobilization of PNs31 with calcium alginate beads was carried out to the design of economic proc
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