Genome mining of cyanide-degrading nitrilases from filamentous fungi
Genome mining of cyanide-degrading nitrilases from filamentous fungi
复制标题
DOI：
10.1007/s00253-008-1559-2
复制
发表时间：
2008-09-01
影响因子：
5
通讯作者：
Benedik, Michael J.
中科院分区：
文献类型：
--
作者：
Basile, Lacy J.;Willson, Richard C.;Benedik, Michael J.
已知多种真菌物种通过氰化物水合酶的作用来降解氰化物，氰化物水合酶是腈酶的一个特殊子集，它将氰化物水解成甲酰胺。在本文中，我们报道了两种以前未知的和未表征的氰化物水化酶从粗神经孢子虫和芽曲霉。利用n端六组氨酸纯化标签克隆了4种氰化物水化酶基因，在大肠杆菌中表达，并利用固定化金属亲和层析技术纯化了4种氰化物水化酶。根据这些酶的相对比活性、pH活性谱、热稳定性和修复镀银和镀铜废水氰化物污染的能力对它们进行了比较。虽然四种氰化物水合酶相似，但粗草氰化物水合酶（CHT）具有最高的热稳定性和最宽的pH范围（bbb50 %活性）。在两种重金属存在的情况下，N. crassa的氰化物降解率也最高。在本研究评价的4种真菌腈酶中，白蜡芽孢杆菌的CHT反应速率最高。这些数据将有助于确定在含氰废物的生物修复中可能使用这些酶的优化程序。与已知的植物病原真菌类似，在KCN存在的情况下，N. crassa和A. nidulans都被诱导表达CHT。
A variety of fungal species are known to degrade cyanide through the action of cyanide hydratases, a specialized subset of nitrilases which hydrolyze cyanide to formamide. In this paper, we report on two previously unknown and uncharacterized cyanide hydratases from Neurospora crassa and Aspergillus nidulans. Recombinant forms of four cyanide hydratases from N. crassa, A. nidulans, Gibberella zeae, and Gloeocercospora sorghi were prepared after their genes were cloned with N-terminal hexahistidine purification tags, expressed in Escherichia coli, and purified using immobilized metal affinity chromatography. These enzymes were compared according to their relative specific activity, pH activity profiles, thermal stability, and ability to remediate cyanide contaminated waste water from silver and copper electroplating baths. Although all four were similar, the N. crassa cyanide hydratase (CHT) has the greatest thermal stability and widest pH range of > 50% activity. N. crassa also demonstrated the highest rate of cyanide degradation in the presence of both heavy metals. The CHT of A. nidulans has the highest reaction rate of the four fungal nitrilases evaluated in this work. These data will help determine optimization procedures for the possible use of these enzymes in the bioremediation of cyanide-containing waste. Similar to known plant pathogenic fungi, both N. crassa and A. nidulans were induced to express CHT by growth in the presence of KCN.