Biotechnology Bulletin ›› 2023, Vol. 39 ›› Issue (4): 103-113.doi: 10.13560/j.cnki.biotech.bull.1985.2022-1270

Previous Articles     Next Articles

Investigation on the Complex Mutagenesis Selection of High-yield Nitrate Reductase Strain Staphylococcus simulans ZSJ6 and Its Enzymatic Properties

ZHAO Sai-sai1(), ZHANG Xiao-dan1, JIA Xiao-yan1, TAO Da-wei1, LIU Ke-yu1, NING Xi-bin1,2,3()   

  1. 1. College of Food Science & Technology, Shanghai Ocean University, Shanghai 201306
    2. Laboratory of Quality & Safety Risk Assessment for Aquatic Products on Storage and Preservation(Shanghai), Ministry of Agriculture and Rural Affairs, Shanghai 201306
    3. National R&D Branch Center for Freshwater Aquatic Products Processing Technology(Shanghai), Shanghai 201306
  • Received:2022-10-17 Online:2023-04-26 Published:2023-05-16

Abstract:

Nitrate reductase can reduce the nitrate in natural meat products to nitrite, avoiding a sudden increase in its content due to direct addition of nitrite and reducing the chance of producing nitrosamines, which is very important for the quality and safety of natural meat products. Staphylococcus simulans D14 screened from sausage as the starting strain, and the different effects of single mutagenesis condition including UV, microwave, and lithium chloride mutagenesis, and two compound mutagenesis methods of UV and microwave combined with lithium chloride on the strains were investigated to obtain high-yield strains of nitrate reductase, and their enzymatic properties were initially explored by shake flask fermentation in order to provide strain selection for fermenters of natural meat products. The optimal conditions for UV, microwave and lithium chloride mutagenesis were determined: UV irradiation for 60 s, microwave irradiation for 80 s and lithium chloride concentration of 1.5%. The mutagenic strain S. simulans ZSJ6 was screened, and the enzyme activity of the strain reached 603.29 U/mg protein, which was 3.59 times higher that of the starting strain, and there was no significant change in the enzyme activity of the mutagenic strain after 8 consecutive passages(P>0.05), indicating that its genetic stability was good. From the results of enzymatic properties, its optimal action temperature was 30℃, the optimal action pH was 7.5, and its stability was excellent, and the enzyme activity retained more than 90% after incubation at pH 7.5 for 2 h. Mg2+, Ca2+, and K+ all promoted enzyme activity, of which Ca2+ had the largest role in promoting the enzyme activity, which was 1.63 times higher than that blank group. Cu2+, Fe2+, Hg2+ and Mn2+ suppressed enzyme activity, among which Cu2+ and Hg2+ had the maximum inhibitory action on the enzyme activity, and the enzyme activity was all inhibited to less than 30%. The above results provide a solution for the transformation of nitrate in natural meat products and the problem of high nitrite concentration brought by the use of nitrite at the initial stage of production, which has research value and potential for application.

Key words: Staphylococcus simulans, nitrate reductase, natural meat products, compound mutagenesis, enzymatic properties, nitrite, nitrate