Enzymatic Characterization of Aminoacylase from Thermophilic Bacteria and Its Application to Amino Acid Production

嗜热细菌氨基酰化酶的酶学表征及其在氨基酸生产中的应用

• 批准号: 61560119
• 负责人: TANIZAWA Katsuyuki
• 金额: $ 1.09万
• 依托单位: Kyoto University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for General Scientific Research (C)
• 财政年份: 1986
• 资助国家: 日本
• 起止时间: 1986 至 1987
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-61560119/
• 关键词: Aminoacylase; Thermostable Enzyme; N-Acylamino Acid; Dipeptidase; Acetamideacrylic Acid; Metallo Enzyme; Enzymatic Analysis; L-フェニルアラニンの酵素生産; 【N!_】-アシルアミノ酸; Aminoacylase; Thermostable Enzyme; N-Acylamino Acid; Dipeptidase; Acetamideacrylic Acid; Metallo Enzyme; Enzymatic Analysis; L-フェニルアラニンの酵素生産; 【N!_】-アシルアミノ酸

This study was undertaken to establish a new rate assay method of aminoacylase, to elucidate enzymological properties of thermostable aminoacylase from thermophilic bacteria to compare with other thermolabile enzymes, and to develop a new enzymatic procedure for the synthesis of amino acids from racemates with the thermostable aminoacylase. The results are summarized as follows:1. A convenient spectrophotometric rate assay for aminoacylase was developped based on the observation that acetamidoacrylate, a synthetic N-acetyl unsaturated amino acid, was hydrolyzed to acetate, ammonia and pyruvate by hog kidney, fungal and bacterial aminoacylases. This assay is linear with time and enzyme concentration, and is useful for kinetic studies of aminoacylase.2. The thermostable aminoacylase was purified to homogeneity from cell extracts of Bacillus thermoglucosidius and characterized. The enzyme has a molecular weight of about 175,000, and is composed of four identical subunits, containing 4 g atoms of zinc per mole of enzyme protein. Great stability at high temperatures and with organic solvents and protein denaturants is a characteristic of the enzyme.3. Enzymological properties of the thermostable dipeptidase from B. stearothermophilus were compared with those of aminoacylase.4. A new enzymatic method for L-phenylalanine production was established.

进行了这项研究是为了建立一种新的氨基酰基酶方法，以阐明嗜热氨基酰基酶从嗜热细菌的酶学特性，以与其他热稳定酶进行比较，并开发一种新的酶促方法，以使氨基酸与热稳定氨基酶合成。结果总结如下：1。根据乙酰氨基丙烯酸酯（一种合成的N-乙酰基氨基酸）的观察，开发了一种方便的分光光度计测定法，将乙酰氨基丙烯酸酯（一种合成的N-乙酰基氨基酸）水解为乙酸，氨和丙酮酸酯，由HOG肾脏，真菌和细菌氨基氨基酶水解。该测定法与时间和酶浓度是线性的，对于氨基酰基酶的动力学研究很有用2。热稳态氨基酰基酶从芽孢杆菌热葡萄糖苷的细胞提取物中纯化为均匀性，并表征。该酶的分子量约为175,000，由四个相同的亚基组成，其中包含每摩尔酶蛋白的4 g原子。高温和有机溶剂和蛋白质变性剂的稳定性是酶的特征。3。将来自硬化芽孢杆菌的热稳态二肽酶的酶学特性与氨基酰基酶的酶学特性进行了比较。4。建立了一种用于L-苯丙氨酸的新酶方法。