Institute of Microbiology, Department of Microbiology , University of Hohenheim , Garbenstrasse 30 , 70599 Stuttgart , Germany.
J Agric Food Chem. 2019 Jan 23;67(3):905-915. doi: 10.1021/acs.jafc.8b04802. Epub 2019 Jan 10.
In the current study, the extracellular endopeptidases from Pseudomonas lundensis and Pseudomonas proteolytica were investigated. The amino acid sequence identity between both endopeptidases is 68%. Both endopeptidases were purified to homogeneity and partially characterized. They were classified as metallopeptidases with a maximum activity at pH 10.0 ( P. lundensis) or 8.5 ( P. proteolytica) at 35 °C. Both remained active in skim milk with 39.7 ± 2.4% and 24.5 ± 3.3%, respectively, of the initial enzyme activity after UHT processing (138 °C for 20 s), indicating the relevance for milk destabilization. The transition points in buffer were determined at 50 °C ( P. lundensis) and 43 °C ( P. proteolytica) using circular dichroism spectroscopy. The loss of the secondary structure at different temperatures was correlated with residual peptidase activities after heat treatment. The ability to destabilize UHT milk was proven by supplementation of skim milk with endopeptidase and storage for 4 weeks.
在本研究中，研究了假单胞菌属lundensis 和假单胞菌属 proteolytica 的细胞外内切肽酶。两种内切肽酶的氨基酸序列同一性为 68%。两种内切肽酶均被纯化至均一性，并进行了部分特性研究。它们被归类为金属肽酶，最适 pH 值为 10.0（P. lundensis）或 8.5（P. proteolytica），在 35°C 下。两种酶在经过 UHT 处理（138°C 20 秒）后，在脱脂乳中仍保持活性，分别具有初始酶活性的 39.7±2.4%和 24.5±3.3%，表明其与牛奶的不稳定性有关。使用圆二色性光谱在缓冲液中确定了转折点，分别为 50°C（P. lundensis）和 43°C（P. proteolytica）。不同温度下二级结构的丧失与热处理后残留的肽酶活性相关。通过在脱脂乳中添加内切肽酶并储存 4 周来证明其使 UHT 牛奶不稳定的能力。
