Discovery of Microbial "Aldoxime-Nitrile Pathway" and Its Application to Nitrile Synthesis

通过使用新的细菌酶“腈水合酶”发现酰胺的积累导致腈水合酶成功地应用于各种酰胺的工业生产。腈水合酶已成为重要的工业酶之一。我们已经成功地在酶促合成腈醛肟通过使用新的微生物酶醛肟脱氢酶，它的工作在新提出的“醛肟腈途径”。(1)醛肟脱氢酶在微生物中的分布相对较广。(2)从芽孢杆菌OxB-1菌株中分离纯化了苯乙醛肟脱氢酶，并克隆了该酶基因，在大肠杆菌中表达。coli JM 109。(3)我们筛选了能将E-吡啶-3-醛肟转化为相应腈的微生物，并从土壤中分离到红球菌YH 3 -3。含有诱导酶的细胞催化醛肟化学计量脱水形成腈。在优化的反应条件下，从10-100 mM醛肟以制备规模合成各种腈。我们还优化了由相应的醛肟通过表达来自芽孢杆菌属菌株OxB-1的苯乙醛肟脱氢酶的大肠杆菌JM 109/poxd 90 F细胞大规模水解芳基腈和烷基腈的条件。我们利用该装置从相应的醛肟出发合成了各种芳腈和烷基腈。(4)从球形红球菌（Rhodococcus globerulus A-4）中分离纯化了醛肟脱氢酶，并对其基因进行了克隆和序列测定。coli JM 109。该酶的基因被证明与腈水合酶基因相连。

Discoveries of an accumulation of amides by the use of new bacterial enzyme "nitrile hydratase" led to the successful application of nitrile hydratase to the industrial production of various amides. Nitrile hydratase e has become one of the most important industrial enzymes. We have been successful in the enzymatic synthesis of nitriles from aldoximes by using the new microbial enzyme aldoxime dehydratase, which works in newly proposed "Aldoxime-Nitrile Pathway".(1) The distribution of aldoxime dehydratase in microorganisms was shown to be relatively wide among microorganisms.(2) Phenylacetaldoxime dehydratase has been purified, characterized and its gene cloned from Bacillus sp. Strain OxB-1 and expressed in E. coli JM 109.(3) We screened for microorganisms to convert E-pyridine-3-aldoxime to the corresponding nitrile and isolated Rhodococcus sp. YH3-3 from soil. The cells containing the inducible enzyme catalyzed the stoichiometric dehydration of aldoximes to form nitriles. Various nitriles were synthesized at preparative scales from 10-100 mM aldoximes under optimized reaction conditions. We also optimized conditions for large-scale synthsis of aryl-and alkyl-nitriles in high yields from the corresponding aldoximes by Escherichia coli JM109/poxd90F cells expressing phenylacetaldoxime dehydratase from Bacillus sp. strain OxB-1. We synthesized various ary1- and alky1-nitriles from the corresponding aldoximes by the transformant cells.(4) Aldoxime dehydratase has been purified, characterized and its gene cloned and sequenced from Rhodococcus globerulus A-4 and expressed in E. coli JM 109. The gene for the enzyme was shown to be linked with the nitrile hydratase gene.