A High-yield Nattokinase Strain: ARTP Mutation Breeding and Medium Optimization

PANDongmei, ZHANGXinqing, YANGChuanlun, TIANJiewei, WANGHongxia, YANGDandan, GUONannan, YUShuaishuai, HANLixia, MAChunfeng, BULiqun, CAIQianqian, GUOZhongrui

PDF(1905 KB)
PDF(1905 KB)
Chinese Agricultural Science Bulletin ›› 2023, Vol. 39 ›› Issue (9) : 115-122. DOI: 10.11924/j.issn.1000-6850.casb2022-0358

A High-yield Nattokinase Strain: ARTP Mutation Breeding and Medium Optimization

Author information +
History +

Abstract

The aim of this paper is to improve the fermentation level of nattokinase and expand its application as a new fibrinolytic drug. A wild strain P producing nattokinase was mutagenized by atmospheric and room temperature plasma (ARTP). A high-yield nattokinase mutant P501 with stable genetic performance was selected and the activity of nattokinase was 1.02 times higher than that of the wild strain. The fermentation medium for nattokinase production was optimized by single factor experiment and response surface experiment. The optimal fermentation medium was as follows: glucose 30 g/L, corn flour 40 g/L, soybean meal 30 g/L, Na2HPO4 3 g/L, K2HPO4 0.3 g/L, MgSO4 0.6 g/L, CaCl2 0.63 g/L, serine 0.07 g/L and soybean oil 0.61 g/L. After verification experiment, the activity of nattokinase reached 9691 Fu/mL, which was 6.68 times higher than that of basic fermentation medium.

Key words

nattokinase / ARTP mutation / single factor experiment / response surface experiment / medium optimization

Cite this article

Download Citations
PAN Dongmei , ZHANG Xinqing , YANG Chuanlun , TIAN Jiewei , WANG Hongxia , YANG Dandan , GUO Nannan , YU Shuaishuai , HAN Lixia , MA Chunfeng , BU Liqun , CAI Qianqian , GUO Zhongrui. A High-yield Nattokinase Strain: ARTP Mutation Breeding and Medium Optimization. Chinese Agricultural Science Bulletin. 2023, 39(9): 115-122 https://doi.org/10.11924/j.issn.1000-6850.casb2022-0358

References

[1]
KOTB E. Activity assessment of microbial fibrinolytic enzymes[J]. Applied microbiology and biotechnology, 2013, 97(15):6647-6665.
Conversion of fibrinogen to fibrin inside blood vessels results in thrombosis, leading to myocardial infarction and other cardiovascular diseases. In general, there are four therapy options: surgical operation, intake of antiplatelets, anticoagulants, or fibrinolytic enzymes. Microbial fibrinolytic enzymes have attracted much more attention than typical thrombolytic agents because of the expensive prices and the side effects of the latter. The fibrinolytic enzymes were successively discovered from different microorganisms, the most important among which is the genus Bacillus. Microbial fibrinolytic enzymes, especially those from food-grade microorganisms, have the potential to be developed as functional food additives and drugs to prevent or cure thrombosis and other related diseases. There are several assay methods for these enzymes; this may due to the insolubility of substrate, fibrin. Existing assay methods can be divided into three major groups. The first group consists of assay of fibrinolytic activity with natural proteins as substrates, e.g., fibrin plate methods. The second and third groups of assays are suitable for kinetic studies and are based on the determination of hydrolysis of synthetic peptide esters. This review will deal primarily with the microorganisms that have been reported in literature to produce fibrinolytic enzymes and the first review discussing the methods used to assay the fibrinolytic activity.
[2]
史旭波, 胡大一. 血栓形成与凝血机制及调节[J]. 临床荟萃, 2007, 22(14):989-991.
[3]
陈伟伟, 高润霖, 刘力生, 等. 《中国心血管病报告 2015》概要[J]. 中国循环杂志, 2015, 31(7):617-622.
[4]
张宝善, 伍晓红, 陈锦屏. 柿单宁研究进展[J]. 陕西师范大学学报:自然科学版, 2008(1):99-105.
[5]
徐君驰, 张青林, 徐莉清, 等. 柿果脱涩机理研究新进展[J]. 园艺学报, 2016, 43(9):1653-1664.
[6]
张海生, 陈锦屏. 柿饼加工中脱涩和反涩机理的研究[J]. 食品工业科技, 2003(12):39-40.
[7]
杜建明. 柿果实的脱涩机理[J]. 食品科学, 1993(4):17-19.
[8]
GUINÉ R P F, BARROCA M J, GONCALVES F J, et al. Artificial neural network modeling of the antioxidant activity and phenolic compounds of bananas submitted to different drying treatments[J]. Food chemistry, 2015, 168:454-459.
[9]
AGHBASHLO M, HOSSEINPOUR S, GHASEMI M. Computer vision technology for real-time food quality assurance during drying process[J]. Trends in food science & technology, 2014, 39(1):76-84.
[10]
张雪, 张晓菲, 王立言, 等. 常压室温等离子体生物诱变育种及其应用研究进展[J]. 化工学报, 2014, 65(7):2676-2684.
大气压射频辉光放电(RF APGD)等离子体具有大气压下操作不需要真空系统、气体温度低、活性粒子浓度高、放电均匀性好、可控性强等特点,能够与各类生物分子发生作用,在生物技术中的应用受到广泛的关注。本研究团队将RF APGD等离子体射流引入生物诱变育种领域,对其物理特性及其与生物大分子和整细胞的作用机制进行了系统研究,并将其开发成新一代高效诱变育种仪,命名为常压室温等离子体(ARTP)诱变育种仪。实践证明,ARTP诱变育种仪具有对操作者安全、环境友好、操作简便、突变快速、突变率高、获得的突变体性状稳定等特点,目前已成功应用于包括细菌、放线菌、真菌、酵母、微藻等在内的四十余种微生物的诱变育种。本文将对ARTP生物育种技术的最新研究进展进行综述,以期ARTP快速生物突变技术在生物进化研究及工业生物菌种改造上发挥重要作用。
[11]
DUAN G W, WU B. Replacing water and nutrients for ethanol production by ARTP derived biogas slurry tolerant Zymomonas mobilis strain[J]. Biotechnology for biofuels, 2019, 12(1):124.
[12]
ZHANG X, ZHANG X F, LI H P, et al. Atmospheric and roomtemperature plasma (ARTP) as a new powerful mutagenesistool[J]. Applied microbiology and biotechnology, 2014, 98(12):5387-5396.
[13]
LAROUSSI M, LEIPOLD F. Evaluation of the roles of reactivespecies, heat, and UV radiation in the inactivation of bacterial cells by air plasmas at atmospheric pressure[J]. International journal of mass spectrometry, 2004, 233(1-3):81-86.
[14]
LI G, LI H P, WANG L Y, et al. Genetic effects of radio-frequency, atmospheric-pressure glow discharges with helium[J]. Applied physics letters, 2008, 92(22):221504.
[15]
崔培梧, 黎继烈, 文蓉, 等. 紫外线诱变青霉P-M1选育柚苷酶高产菌株研究[J]. 北方园艺, 2009(2):42-44.
[16]
WANG H, SUN X, WANG L, et al. Coproduction of menaquinone-7 and nattokinase by Bacillus subtilis using soybean curd residue as a renewable substrate combined with a dissolved oxygen control strategy[J]. Annals of microbiology, 2018, 68(10):655-665.
[17]
李勇昊, 姜永生, 周长海, 等. 响应面法优化里氏木霉RutC-30产纤维素酶液体培养基[J]. 中国酿造, 2012(4):29-32.
[18]
高泽鑫. 高产纳豆激酶菌株的筛选及其酶学稳定性的研究[D]. 贵阳: 贵州大学, 2018.
[19]
满丽莉, 向殿军. 传统发酵豆制品中纳豆激酶产生菌的筛选及发酵培养基的优化[J]. 中国调味品, 2019, 44(6):44-49,55.
[20]
张海粟, 王家林, 于江淼. 纳豆激酶的研究及展望[J]. 食品与发酵科技, 2019, 55(4):92-95,111.
Share on Mendeley
PDF(1905 KB)

Collection(s)

Triticum aestivum L.

Accesses

Citation

Detail

Sections
Recommended

/