Obtaining Efficient Mutant from the Wild Type Bacillus subtilis E6-5 by Physical and Chemical Mutagenesis for High Efficiency Protease Production, Optimizing the Mutant's Culture Medium

IF 1 Q3 MULTIDISCIPLINARY SCIENCES gazi university journal of science Pub Date : 2023-04-11 DOI:10.35378/gujs.1191006
Büşra Özalpar, E. Demirkan, Tuba Sevgi̇
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Abstract

In this study, to enhance protease production, the wild type of Bacillus subtilis E6-5 was mutagenized by random mutagenesis using ultraviolet radiation and ethidium bromide. After combined treatment, several mutants were obtained. Among these mutants, the mutant strain with the largest proteolytic zone diameter (25 mm) was selected and named Bacillus subtilis ATA38. The enzyme production capacity of the obtained mutant was tested and the mutant strain (404 IU/mL at 24 hours) produced 6.7 times more enzyme than the parental strain (60 IU/mL at 32 hours). The effects of some important parameters in the growth medium on enzyme production were examined. The best carbon, organic nitrogen and metal ion were obtained with wheat starch (525 IU/mL), meat extract (850 IU/mL) and KCl+CaCl2 (548 IU/mL), respectively. pH 6.0, 37°C, 200 rpm, inoculum age 18 hours and inoculation amount 1% were obtained as the best physical factors.To further increase the yield, the best nutritional and physical parameters were combined to create a new modified medium. It was determined that the enzyme yield with mutant strain increased 2.7 times in the modified medium (1096 IU/mL) compared to the control (404 U/mL). The mutant strain (1096 IU/mL) showed an 18.2-fold increase in production compared to the wild type (60 IU/mL) in the modified medium. Protease enzyme obtained from ATA38 mutant strain may have great potential in industry for different purposes.
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通过物理和化学诱变获得野生型枯草芽孢杆菌E6-5高效蛋白酶突变体,并优化突变体培养基
本研究利用紫外辐射和溴化乙啶对野生型枯草芽孢杆菌E6-5进行随机诱变,以提高蛋白酶的产量。综合处理后,获得了多个突变体。在这些突变体中,选择了蛋白水解带直径最大(25 mm)的突变株,命名为枯草芽孢杆菌ATA38。对获得的突变株产酶能力进行了测试,结果表明,突变株(24小时404 IU/mL)的产酶量是亲本菌株(32小时60 IU/mL)的6.7倍。考察了培养基中一些重要参数对产酶的影响。以小麦淀粉(525 IU/mL)、肉精(850 IU/mL)和KCl+CaCl2 (548 IU/mL)分别获得最佳碳、有机氮和金属离子。最佳物理条件为pH 6.0, 37℃,200 rpm,接种时间18 h,接种量1%。为了进一步提高产量,将最佳的营养参数和物理参数结合起来,创造了一种新的改良培养基。结果表明,在改良培养基(1096 IU/mL)中,突变菌株的酶产率比对照(404 U/mL)提高了2.7倍。与野生型(60 IU/mL)相比,突变菌株(1096 IU/mL)在改良培养基中的产量增加了18.2倍。从ATA38突变株中获得的蛋白酶在工业上具有很大的应用潜力。
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来源期刊
gazi university journal of science
gazi university journal of science MULTIDISCIPLINARY SCIENCES-
CiteScore
1.60
自引率
11.10%
发文量
87
期刊介绍: The scope of the “Gazi University Journal of Science” comprises such as original research on all aspects of basic science, engineering and technology. Original research results, scientific reviews and short communication notes in various fields of science and technology are considered for publication. The publication language of the journal is English. Manuscripts previously published in another journal are not accepted. Manuscripts with a suitable balance of practice and theory are preferred. A review article is expected to give in-depth information and satisfying evaluation of a specific scientific or technologic subject, supported with an extensive list of sources. Short communication notes prepared by researchers who would like to share the first outcomes of their on-going, original research work are welcome.
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