产氨肽酶芽孢菌的筛选及酶学性质研究
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(1.安徽省农业科学院农产品加工研究所 合肥 230031;2.安徽省食品微生物发酵与功能应用工程实验室 合肥 230031)

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科技部重点专项(SQ2020YFF0404523);安徽省重大科技专项(202003b06020027)


Screening of a Marine Aminopeptidase Producing Bacterium and Its Enzymatic Properties
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(1.Institute of Agricultural Products Processing, Anhui Academy of Agricultural Sciences, Hefei 230031;2.Anhui Engineering Laboratory of Microbial Fermentation and Functional Application of Food, Hefei 230031)

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    摘要:

    氨肽酶作为一种蛋白质水解酶被广泛应用于食品发酵加工领域。筛选高产氨肽酶菌株,并对菌株中风味酶的酶学性质进行研究,对开发高质量豆制品发酵剂具有重要意义。本文利用以L-亮氨酸-对硝基苯胺(L-Leu-PNA)为底物的分离培养基筛选出1株产氨肽酶的芽孢菌XYY1,通过形态特征观察、生理生化试验和16S rDNA技术对其进行鉴定,采用单因素实验和响应面方法优化产酶培养条件,并对菌株XYY1所产氨肽酶Lap-Y的酶学性质进行研究。结果表明,菌株XYY1经鉴定为地衣芽孢杆菌,起始pH值为6.5,置36 ℃条件下恒温培养35.5 h时,氨肽酶含量高达(4 132.75±11.94)U。酶学性质研究表明反应温度为55 ℃时Lap-Y的相对活性最高,且热稳定性良好;最适pH值为9.0,在中性偏碱环境下稳定性较好,属于偏碱性酶;金属离子Mg2+、Fe2+、CO2+和Ca2+可激活Lap-Y活性,其中Mg2+为1.0 mmol/L时对氨肽酶Lap-Y的激活作用最显著,乙二胺四乙酸(EDTA)、二硫苏糖醇(DTT)、十二烷基硫酸钠(SDS)对Lap-Y都有较强的抑制作用。地衣芽孢杆菌XYY1所产的氨肽酶Lap-Y适合工业化生产,对食品、医疗等行业具有极高的应用价值。

    Abstract:

    Aminopeptidase as a protein hydrolase is widely used in food fermentation and processing. Screening high aminopeptidase-producing strains and studying the enzymatic properties of the flavor enzymes in the strains are important for the development of high-quality soybean product ferments. In this paper, an aminopeptidase-producing strain of Bacillus sp. XYY1 was screened using L-Leu-PNA as substrate in the isolation medium, and identified by morphological observation, physiological and biochemical tests, and 16S rDNA technology, and optimized the enzyme-producing culture conditions by using a one-factor experiment and response surface methodology, and studied the enzymatic properties of the aminopeptidase Lap-Y produced by strain XYY1. The enzyme properties of the aminopeptidase Lap-Y produced by strain XYY1 were investigated. The results showed that strain XYY1 was identified as Bacillus licheniformis, with a starting pH of 6.5, and the content of aminopeptidase was as high as(4,132.75±11.94) U when it was cultured at a constant temperature of 36 ℃ for 35.5 h. The study of the enzymatic properties showed that the relative activity of Lap-Y was highest at the reaction temperature of 55 ℃ with good thermal stability, and that the optimal pH was 9.0, which was good for enzyme stability in the neutral alkaline environment, which was alkaline. The optimum pH value was 9.0, and the enzyme was alkaline in a neutral alkaline environment. Metal ions Mg2+, Fe2+, CO2+ and Ca2+ could activate the activity of Lap-Y, among which the activation effect of Mg2+ was the most significant for the aminopeptidase Lap-Y when Mg2+ was 1.0 mmol/L, and the activation effect of EDTA, dithiothreitol (DTT) and sodium dodecyl sulphate (SDS) had a stronger inhibitory effect on the activity of Lap-Y. inhibitory effect. The aminopeptidase Lap-Y produced by Bacillus licheniformis XYY1 is suitable for industrialized production and has high application value for food and medical industries.

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李冬琪,程江华,万娅琼,徐雅芫.产氨肽酶芽孢菌的筛选及酶学性质研究[J].中国食品学报,2024,24(11):71-85

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  • 收稿日期:2023-11-19
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  • 在线发布日期: 2024-12-25
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