Food Biotechnology Laboratory, BOKU-University of Natural Resources and Life Sciences Vienna, Muthgasse 11, Vienna 1190, Austria.
Biomolecules. 2013 Aug 16;3(3):535-52. doi: 10.3390/biom3030535.
Pyranose dehydrogenase (PDH) is a flavin-dependent sugar oxidoreductase that is limited to a rather small group of litter-degrading basidiomycetes. The enzyme is unable to utilize oxygen as an electron acceptor, using substituted benzoquinones and (organo) metal ions instead. PDH displays a broad substrate specificity and intriguing variations in regioselectivity, depending on substrate, enzyme source and reaction conditions. In contrast to the related enzyme pyranose 2-oxidase (POx), PDHs from several sources are capable of oxidizing α- or β-1→4-linked di- and oligosaccharides, including lactose. PDH from A. xanthoderma is able to perform C-1 and C-2 oxidation, producing, in addition to lactobionic acid, 2-dehydrolactose, an intermediate for the production of lactulose, whereas PDH from A. campestris oxidizes lactose nearly exclusively at the C-1 position. In this work, we present the isolation of PDH-encoding genes from A. campestris (Ac) and A. xanthoderma (Ax) and a comparison of other so far isolated PDH-sequences. Secretory overexpression of both enzymes in Pichia pastoris was successful when using their native signal sequences with yields of 371 U·L-1 for AxPDH and 35 U·L-1 for AcPDH. The pure enzymes were characterized biochemically and tested for applications in carbohydrate conversion reactions of industrial relevance.
吡喃糖脱氢酶（PDH）是一种黄素依赖性糖氧化还原酶，仅限于一小部分分解落叶的担子菌。该酶不能将氧气用作电子受体，而是使用取代的苯醌和（有机）金属离子。PDH 显示出广泛的底物特异性和有趣的区域选择性变化，具体取决于底物、酶源和反应条件。与相关酶吡喃糖 2-氧化酶（POx）不同，来自几种来源的 PDH 能够氧化 α-或β-1→4 键合的二糖和寡糖，包括乳糖。黄孢原毛平革菌的 PDH 能够进行 C-1 和 C-2 氧化，除了产生乳酮酸外，还产生 2-脱水乳糖，这是乳果糖生产的中间产物，而来自野油菜黄单胞菌的 PDH 几乎仅在 C-1 位置氧化乳糖。在这项工作中，我们从野油菜黄单胞菌（Ac）和黄孢原毛平革菌（Ax）中分离出编码 PDH 的基因，并比较了其他迄今为止分离出的 PDH 序列。当使用其天然信号序列时，毕赤酵母中两种酶的分泌过表达都是成功的，AxPDH 的产率为 371 U·L-1，AcPDH 的产率为 35 U·L-1。纯酶的生化特性得到了表征，并测试了它们在具有工业相关性的碳水化合物转化反应中的应用。
