Aromatic nitriles are important structural motifs that frequently existed in pharmaceutical drugs. Due to the convenient synthesis of aldoximes from aldehydes, the dehydration of aldoximes to corresponding nitriles by aldoxime dehydratases (Oxds) is considered as a safe and robust enzymatic production route. Although the Oxd genes are widely distributed in microbial kingdom, so far less than ten Oxds were expressed and further characterized. In this study, we found 26 predicted putative Oxd genes from the GenBank database using a genome mining strategy. The Oxd gene from Pseudomonas putida F1 was cloned and functionally expressed in Escherichia coli BL21 (DE3). The amino acid sequence of OxdF1 shows high identities of 33∼85 % to other characterized Oxds, and contained a ferrous heme as the catalytic site. The optimum reaction pH and temperature of recombinant OxdF1 were 7.0 and 35 °C, respectively. OxdF1 was stable in pH 7.0 potassium phosphate buffer at 30 °C, and its half-life was approximately 3.8 h. OxdF1 can efficiently dehydrate aromatic and heterocyclic aldoximes to nitriles, such as 2-bromobenzaldoxime, 2-chloro-6-fluorobenzaldoxime, thiophene-2-carboxaldoxime, and pyridine-3-aldoxime. Therefore, the recombinant OxdF1 shows a potential application in the cyanide-free synthesis of aromatic nitriles.

芳香腈是药物中经常存在的重要结构基序。由于醛肟的方便合成，醛肟脱水酶（Oxds）将醛肟脱水为相应的腈被认为是一种安全且稳健的酶促生产途径。尽管 Oxd 基因在微生物界广泛分布，但迄今为止表达和进一步表征的 Oxd 不到 10 个。在这项研究中，我们使用基因组挖掘策略从 GenBank 数据库中发现了 26 个预测的假定 Oxd 基因。恶臭假单胞菌F1的 Oxd 基因在大肠杆菌中被克隆和功能表达BL21 (DE3)。OxdF1 的氨基酸序列与其他具有特征的 Oxds 具有 33~85% 的高度同一性，并含有一个亚铁血红素作为催化位点。重组OxdF1的最佳反应pH和温度分别为7.0和35°C。OxdF1 在 30 °C 的 pH 7.0 磷酸钾缓冲液中稳定，其半衰期约为 3.8 h。OxdF1 可以有效地将芳香族和杂环醛肟脱水成腈，例如 2-溴苯甲醛肟、2-氯-6-氟苯甲醛肟、噻吩-2-羧醛肟和吡啶-3-醛肟。因此，重组 OxdF1 在芳香腈的无氰合成中显示出潜在的应用。