Kettner Lucas, Freund Alexander, Bechtel Anna, Costa-Catala Judit, Fischer Lutz
Department of Biotechnology and Enzyme Science, Institute of Food Science and Biotechnology, University of Hohenheim, Garbenstr. 25, 70599 Stuttgart, Germany.
Departament de Nutrició, Ciències de l'Alimentació i Gastronomia, Campus de l'Alimentació de Torribera, Universitat de Barcelona, Av. Prat de la Riba 171, 08921 Santa Coloma de Gramenet, Spain.
Foods. 2025 Sep 3;14(17):3093. doi: 10.3390/foods14173093.
A novel diamine oxidase (DAO) was discovered in the bacterium (DAO-GH). The gene of DAO-GH was integrated into the genome of the yeast and recombinantly produced under control of the methanol-inducible AOX1 promoter in a bioreactor cultivation. A high DAO activity of 70.2 ± 5.2 µkat/L (5.25 ± 0.22 µkat/g) was yielded after 90 h of cultivation. The DAO-GH was partially purified by the polyethyleneimine precipitation of nucleic acids, fractionated ammonium sulfate precipitation and hydrophobic interaction chromatography, resulting in a specific DAO activity of 19.7 µkat/g. The DAO-GH was then biochemically investigated regarding its potential for histamine and tyramine degradation in fermented foods and the human small intestine. Interestingly, the DAO-GH showed activity even at a low pH of 5 and low temperature of 6 °C. Both histamine and tyramine were effectively degraded and DAO-GH showed especially very high affinity towards tyramine ( of 0.009 mM). The DAO-GH was shown to be capable of degrading around 20% of the initially applied histamine in tuna paste (pH 5.6) at 5 °C within 24 h and completely degraded the histamine in a simulated intestinal fluid within 1.5 h in bioconversion experiments. The DAO-GH was spray-dried for the production of a storable enzyme preparation. Only around 17% of activity were lost in this process and the DAO-GH remained stable at room temperature for at least 3 months. The discovery of this DAO with its very advantageous biochemical properties allows the preparation of histamine-reduced or -free fermented foods by a simple enzymatic treatment or the treatment of histamine intolerance symptoms as a dietary supplement or medicine.
在细菌中发现了一种新型二胺氧化酶（DAO-GH）。将DAO-GH的基因整合到酵母基因组中，并在生物反应器培养中在甲醇诱导型AOX1启动子的控制下进行重组生产。培养90小时后，产生了70.2±5.2μkat/L（5.25±0.22μkat/g）的高DAO活性。通过核酸的聚乙烯亚胺沉淀、分级硫酸铵沉淀和疏水相互作用色谱法对DAO-GH进行部分纯化，得到的特定DAO活性为19.7μkat/g。然后对DAO-GH进行了生化研究，考察其在发酵食品和人类小肠中降解组胺和酪胺的潜力。有趣的是，DAO-GH即使在低pH值5和低温6℃下也具有活性。组胺和酪胺均被有效降解，并且DAO-GH对酪胺表现出特别高的亲和力（解离常数为0.009 mM）。在生物转化实验中，DAO-GH在5℃下24小时内能够降解金枪鱼酱（pH 5.6）中约20%的初始添加组胺，并在1.5小时内将模拟肠液中的组胺完全降解。为了生产可储存的酶制剂，对DAO-GH进行了喷雾干燥。在此过程中仅损失了约17%的活性，并且DAO-GH在室温下至少3个月保持稳定。这种具有非常有利生化特性的DAO的发现，使得通过简单的酶处理制备组胺含量降低或不含组胺的发酵食品，或者作为膳食补充剂或药物治疗组胺不耐受症状成为可能。
