Structural diversification during glucosinolate breakdown: mechanisms of thiocyanate, epithionitrile and simple nitrile formation
Structural diversification during glucosinolate breakdown: mechanisms of thiocyanate, epithionitrile and simple nitrile formation
复制标题
DOI：
10.1111/tpj.14327
复制
发表时间：
2019-07-01
期刊：
影响因子：
7.2
通讯作者：
Brand, Wolfgang
中科院分区：
文献类型：
--
作者：
Eisenschmidt-Boenn, Daniela;Schneegans, Nicola;Brand, Wolfgang
次生代谢的特点是结构多样性。在这里，我们已经解决了潜在的机制结构多样化后，损伤诱导的硫代葡萄糖苷，一组硫代葡萄糖苷中发现的拟南芥。芥子油苷活化的经典途径包括黑芥子酶催化的水解和糖苷配基重排为异硫氰酸酯。菊科植物具有特异性蛋白，即非血红素铁蛋白，其通过未知机制干扰该反应来促进替代产物的形成。我们已经使用了结构信息的硫氰酸盐形成蛋白质从Thlaspi arvense（TaTFP），测试的影响，突出的环在其β-螺旋桨结构的一侧上的产品形成使用烯丙基硫代葡萄糖苷配基作为底物。在硅环结构采样和半经验量子力学计算确定了一个3L 2环构象，使铁2+辅因子与烯丙基侧链的双键相互作用。只有这种安排，使烯丙基硫氰酸盐，一个特定的产品的TaTFP的形成。模拟3，4-epithiobutane腈的形成，第二个已知的产品的TaTFP，需要一个替代的基板对接安排，其中Fe 2+与苷元硫醇盐相互作用。与这些结果相一致，参与构象变化的3L 2氨基酸残基的取代以及相邻环的关键氨基酸残基的交换影响了在TaTFP体外存在下黑芥子酶催化的烯丙基芥子油苷水解后获得的烯丙基硫氰酸酯与环硫腈的比例。基于这些见解，我们建议，specifier蛋白是催化剂，可能被归类为Fe 2 +-依赖的lyases.Significance声明硫代葡萄糖苷分解后的结构多样化是由specifier蛋白，一组非血红素铁蛋白控制。我们的研究提供了机制的见解，确定指定的蛋白作为Fe 2+依赖的C-S/C-C裂解酶。与简单的腈形成过程中Fe 2+在硫提取中的作用相比，硫氰酸盐和环硫腈的形成依赖于Fe 2 +/Fe 3+作为氧化还原伙伴。Thlaspi arvense硫氰酸盐形成蛋白的硫氰酸盐形成需要构象变化，从而能够实现替代底物对接姿势。
Secondary metabolism is characterized by an impressive structural diversity. Here, we have addressed the mechanisms underlying structural diversification upon damage-induced activation of glucosinolates, a group of thioglucosides found in the Brassicales. The classical pathway of glucosinolate activation involves myrosinase-catalyzed hydrolysis and rearrangement of the aglucone to an isothiocyanate. Plants of the Brassicaceae possess specifier proteins, i.e. non-heme iron proteins that promote the formation of alternative products by interfering with this reaction through unknown mechanisms. We have used structural information available for the thiocyanate-forming protein from Thlaspi arvense (TaTFP), to test the impact of loops protruding at one side of its beta-propeller structure on product formation using the allylglucosinolate aglucone as substrate. In silico loop structure sampling and semiempirical quantum mechanical calculations identified a 3L2 loop conformation that enabled the Fe2+ cofactor to interact with the double bond of the allyl side chain. Only this arrangement enabled the formation of allylthiocyanate, a specific product of TaTFP. Simulation of 3,4-epithiobutane nitrile formation, the second known product of TaTFP, required an alternative substrate docking arrangement in which Fe2+ interacts with the aglucone thiolate. In agreement with these results, substitution of 3L2 amino acid residues involved in the conformational change as well as exchange of critical amino acid residues of neighboring loops affected the allylthiocyanate versus epithionitrile proportion obtained upon myrosinase-catalyzed allylglucosinolate hydrolysis in the presence of TaTFP invitro. Based on these insights, we propose that specifier proteins are catalysts that might be classified as Fe2+-dependent lyases.Significance Statement Structural diversification upon glucosinolate breakdown is controlled by specifier proteins, a group of non-heme iron proteins. Our study provides mechanistic insights that identify specifier proteins as Fe2+-dependent C-S/C-C lyases. By contrast with the role of Fe2+ in sulfur abstraction during simple nitrile formation, thiocyanate and epithionitrile formation depends on Fe2+/Fe3+ as a redox partner. Thiocyanate formation by Thlaspi arvense thiocyanate-forming protein requires a conformational change that enables an alternative substrate docking pose.