Cage escape competes with geminate recombination during alkane hydroxylation by the diiron oxygenase AlkB
Cage escape competes with geminate recombination during alkane hydroxylation by the diiron oxygenase AlkB
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DOI：
10.1002/anie.200801184
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发表时间：
2008-01-01
影响因子：
16.6
通讯作者：
Groves, John T.
中科院分区：
文献类型：
--
作者：
Austin, Rachel N.;Luddy, Kate;Groves, John T.
恶臭假单胞菌GPo 1的烷烃羟化酶AlkB是一类典型的跨膜二铁加氧酶，其催化羟基化、环氧化和去饱和反应。[1，2]这些酶由于其对全球碳氢化合物代谢的影响而引起了相当大的兴趣，[3]它们在实际生物催化应用中的潜力，以及由此产生的合成仿生催化剂设计的灵感。[4]虽然AlkB或任何密切相关的蛋白质的三维结构是未知的，拓扑建模预测的结构包括六个跨膜螺旋与催化铁二联体附加到螺旋束的细胞质末端。[5]穆斯堡尔数据和丙氨酸扫描表明，二铁结合位点富含组氨酸，[6]与在血红蛋白中发现的一样，并且与在二铁羟化酶sMMO [7]和T4 MOh中发现的主要羧酸结合基序相反。[8]通过蛋白质侧链突变，已经确定了一个长的，疏水性的底物结合通道内的束，是调整接受中等长度的烷烃。[5]AlkB是第一个烷烃羟化酶，催化过程中产生一个长寿命的底物碳自由基，所揭示的诊断骨架重排的烃探针降冰片烷。[9]在本文中，我们报告了使用一组自由基时钟底物的AlkB羟基化反应的结果，这些底物显示出跨越五个数量级的固有重排速率，从双环-[3.1]的适度缓慢的2.8 × 107 sq-1。0]己烷[11]转化为超快1011 sample 1，用于反式-1-甲基-2-
The alkane hydroxylase AlkB of Pseudomonas putida GPo1 is typical of a large class of membrane-spanning diiron oxygenases that catalyze hydroxylation, epoxidation, and desaturation reactions.[1, 2] These enzymes are of considerable interest due to their impact on global hydrocarbon metabolism,[3] their potential for practical biocatalytic application, and the resulting inspiration for the design of synthetic biomimetic catalysts.[4] Although the three-dimensional structures of AlkB or any closely related proteins are unknown, topology modeling has predicted a structure comprised of six membrane-spanning helices with the catalytic iron diad appended to the cytoplasmic termini of the helix bundle.[5] Mössbauer data and alanine scanning have suggested that the diiron binding site is histidine-rich,[6] as found in hemerythrin, and in contrast to the predominantly carboxylate binding motifs found in the diiron hydroxylases sMMO [7] and T4MOh.[8] Through protein side-chain mutations, a long, hydrophobic substrate-binding channel within the bundle has been identified that is tuned to accept medium-length alkanes.[5] AlkB was the first alkane hydroxylase shown to generate a longlived substrate carbon radical during catalysis, as revealed by diagnostic skeletal rearrangements of the hydrocarbon probe norcarane.[9]Herein we report results for the AlkB hydroxylation reaction using a panel of radical-clock substrates that display intrinsic rearrangement rates spanning five orders of magnitude, from a moderately slow 2.8 107 sÀ1 for bicyclo-[3.1. 0] hexane [11] to an ultrafast 1011 sÀ1 for trans-1-methyl-2-