The crystal structure of the ivy Δ4-16:0-ACP desaturase reveals structural details of the oxidized active site and potential determinants of regioselectivity
The crystal structure of the ivy Δ4-16:0-ACP desaturase reveals structural details of the oxidized active site and potential determinants of regioselectivity
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DOI：
10.1074/jbc.m702520200
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发表时间：
2007-07-06
影响因子：
4.8
通讯作者：
Shanklin, John
中科院分区：
文献类型：
--
作者：
Guy, Jodie E.;Whittle, Edward;Shanklin, John
来自常春藤（Hedera helix）（英国常春藤（English ivy））的多功能酰基-酰基载体蛋白（ACP）去饱和酶催化16：0-ACP的Delta（4）去饱和和18：0-ACP的Delta（9）去饱和，并进一步将Delta（9）-16：1或Delta（9）-18：1去饱和为相应的Delta（4，9）二烯。该酶的晶体结构已经被解析到1.95埃分辨率，并且类似于3.2埃的铁-铁距离和μ-氧桥的存在都揭示了这是氧化的FeIII-FeIII形式的去饱和酶的唯一报道的结构。在蓖麻去饱和酶的氧化活性位点和还原活性位点之间观察到显著差异; His（227）与Fe 2的配位丢失，并且在还原结构中桥接两个铁离子的Glu（224）的侧链不与任一铁相互作用。虽然已经观察到其他二铁蛋白氧化的羧酸盐位移，但这是第一个超出两个铁离子配位范围的残基。常春藤和蓖麻结构的比较显示表面氨基酸接近环的基板结合腔和其他内衬的腔的下部，是其独特的基板特异性的潜在决定因素。我们提出了一个假设，侧链包装的差异解释了明显的悖论，即几个残基内衬的空腔中的常春藤去饱和酶的较低的部分是笨重的比他们的等价物在蓖麻酶，尽管常春藤酶的必要性，以容纳三个以上的碳超出二铁网站。
The multifunctional acyl-acyl carrier protein (ACP) desaturase from Hedera helix ( English ivy) catalyzes the Delta(4) desaturation of 16:0-ACP and the Delta(9) desaturation of 18: 0-ACP and further desaturates Delta(9)-16: 1 or Delta(9)-18: 1 to the corresponding Delta(4,9) dienes. The crystal structure of the enzyme has been solved to 1.95 angstrom resolution, and both the iron-iron distance of similar to 3.2 angstrom and the presence of a mu-oxo bridge reveal this to be the only reported structure of a desaturase in the oxidized FeIII-FeIII form. Significant differences are seen between the oxidized active site and the reduced active site of the Ricinus communis ( castor) desaturase; His(227) coordination to Fe2 is lost, and the side chain of Glu(224), which bridges the two iron ions in the reduced structure, does not interact with either iron. Although carboxylate shifts have been observed on oxidation of other diiron proteins, this is the first example of the residue moving beyond the coordination range of both iron ions. Comparison of the ivy and castor structures reveal surface amino acids close to the annulus of the substrate-binding cavity and others lining the lower portion of the cavity that are potential determinants of their distinct substrate specificities. We propose a hypothesis that differences in side chain packing explains the apparent paradox that several residues lining the lower portion of the cavity in the ivy desaturase are bulkier than their equivalents in the castor enzyme despite the necessity for the ivy enzyme to accommodate three more carbons beyond the diiron site.