Kinetic characterisation of the FAD dependent monooxygenase TropB and investigation of its biotransformation potential
Kinetic characterisation of the FAD dependent monooxygenase TropB and investigation of its biotransformation potential
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DOI：
10.1039/c5ra06693j
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发表时间：
2015-01-01
期刊：
影响因子：
3.9
通讯作者：
Cox, Russell J.
中科院分区：
文献类型：
--
作者：
Abood, Amira;Al-Fahad, Ahmed;Cox, Russell J.
实现芳香族化合物的区域特异性羟基化仍然是合成化学中的主要挑战。相比之下，这种转化在自然界中通过FAD依赖性单加氧酶的作用容易地完成。在这里，我们报告的动力学特性的一个这样的酶，TropB，从柄基酸生物合成途径。类似物的TropB天然底物，3-甲基-orcinaldehyde，合成和用于检查这种酶的底物选择性。TropB显示出广泛的底物耐受性，例如接受含有一系列具有不同电子和空间性质的C-1取代基的单环芳族底物。这些基团包括硝基、亚硝酰基、烷基和芳基酮基。然而，双环底物被TropB拒绝。此外，C-5取代基上的单环芳族底物是不容忍的，而6-甲基基团的存在下被发现是重要的底物结合。采用对接研究来调查和理解TropB的广泛底物选择性并鉴定其底物的关键结构元件。我们的工作表明，TropB是一个有吸引力的目标，生物催化剂工程和工业芳族羟基化。
Achieving regio-specific hydroxylation of aromatic compounds remains a major challenge in synthetic chemistry. By contrast, this transformation is readily accomplished in nature through the action of FAD-dependant monooxygenase enzymes. Here, we report the kinetic characterisation of one such enzyme, TropB, from the stipitatic acid biosynthetic pathway. Analogues of the TropB natural substrate, 3-methyl-orcinaldehyde, were synthesised and used to examine the substrate selectivity of this enzyme. TropB displays broad substrate tolerance, for instance accepting single-ring aromatic substrates containing a range of C-1 substituents with varying electronic and steric properties. These include nitro, nitrosyl, alkyl, and aryl keto groups. Bicyclic substrates, however, were rejected by TropB. Additionally, C-5 substituents on single-ring aromatic substrates were not tolerated whereas the presence of a 6-methyl group was found to be important for substrate binding. Docking studies were employed to investigate and understand the broad substrate selectivity of TropB and identifies the key structural elements of its substrates. Our work has shown that TropB is an attractive target for biocatalyst engineering and industrial aromatic hydroxylation.