The Cytochrome P450 OxyA from the Kistamicin Biosynthesis Cyclization Cascade is Highly Sensitive to Oxidative Damage.
The Cytochrome P450 OxyA from the Kistamicin Biosynthesis Cyclization Cascade is Highly Sensitive to Oxidative Damage.
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DOI：
10.3389/fchem.2022.868240
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发表时间：
2022
影响因子：
5.5
通讯作者：
Cryle, Max J.
中科院分区：
文献类型：
--
作者：
Greule, Anja;Izore, Thierry;Machell, Daniel;Hansen, Mathias H.;Schoppet, Melanie;De Voss, James J.;Charkoudian, Louise K.;Schittenhelm, Ralf B.;Harmer, Jeffrey R.;Cryle, Max J.
细胞色素P450酶（P450）是一个单加氧酶超家族，利用半胱氨酸硫酸连接的血红素片段进行广泛的氧化转化。鉴于p450酶在其活性循环中形成的活性中间体的氧化能力，值得注意的是，这些酶可以避免自氧化，并将轴向半胱氨酸配体保持在去质子化（因此高度酸性）的硫酸盐形式。虽然对P450折叠过程中血红素结合的过程知之甚少，但在P450活性位点内，有一种血红素取向的压倒性偏好。事实上，迄今为止，很少有结构包含替代血红素取向，其中两个是来自糖肽抗生素（GPA）生物合成的OxyA同源物。鉴于OxyA酶对不寻常的血红素取向的明显偏好，我们研究了来自kisistamicin生物合成（OxyAkis）的OxyA同源物，这是一种非典型的GPA。我们确定OxyAkis对过氧化氧化物的氧化损伤高度敏感，紫外和EPR测量显示血红素信号的快速漂白。我们确定了OxyAkis的结构，并发现在这种酶中存在血红素取向的混合种群。我们的分析进一步揭示了血红素片段的可能修饰，这只存在于蛋白质中存在替代血红素取向的样品中。这些结果表明，在P450催化过程中，细胞色素P450中典型的血红素取向可以帮助防止对血红素的潜在损伤，从而防止酶的失活。这也表明一些参与GPA生物合成的P450酶可能特别容易受到氧化损伤，因为在它们的活性位点发现了血红素取向。
Cytochrome P450 enzymes (P450s) are a superfamily of monooxygenases that utilize a cysteine thiolate–ligated heme moiety to perform a wide range of demanding oxidative transformations. Given the oxidative power of the active intermediate formed within P450s during their active cycle, it is remarkable that these enzymes can avoid auto-oxidation and retain the axial cysteine ligand in the deprotonated—and thus highly acidic—thiolate form. While little is known about the process of heme incorporation during P450 folding, there is an overwhelming preference for one heme orientation within the P450 active site. Indeed, very few structures to date contain an alternate heme orientation, of which two are OxyA homologs from glycopeptide antibiotic (GPA) biosynthesis. Given the apparent preference for the unusual heme orientation shown by OxyA enzymes, we investigated the OxyA homolog from kistamicin biosynthesis (OxyAkis), which is an atypical GPA. We determined that OxyAkis is highly sensitive to oxidative damage by peroxide, with both UV and EPR measurements showing rapid bleaching of the heme signal. We determined the structure of OxyAkis and found a mixed population of heme orientations present in this enzyme. Our analysis further revealed the possible modification of the heme moiety, which was only present in samples where the alternate heme orientation was present in the protein. These results suggest that the typical heme orientation in cytochrome P450s can help prevent potential damage to the heme—and hence deactivation of the enzyme—during P450 catalysis. It also suggests that some P450 enzymes involved in GPA biosynthesis may be especially prone to oxidative damage due to the heme orientation found in their active sites.
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影响因子：
14.8
作者：
Kelley LA;Mezulis S;Yates CM;Wass MN;Sternberg MJ
通讯作者：
Sternberg MJ
影响因子：
5.2
作者：
Brieke, Clara;Cryle, Max J.
通讯作者：
Cryle, Max J.
DOI：
10.1107/s0907444904019158
发表时间：
2004-12-01
影响因子：
2.2
作者：
Emsley, P;Cowtan, K
通讯作者：
Cowtan, K
影响因子：
3.9
作者：
Cryle, Max J.;Staaden, Jessica;Schlichting, Ilme
通讯作者：
Schlichting, Ilme
影响因子：
3.2
作者：
Haslinger, Kristina;Maximowitsch, Egle;Cryle, Max J.
通讯作者：
Cryle, Max J.