Distinct structural elements dictate the specificity of the type III pentaketide synthase from Neurospora crassa.
Distinct structural elements dictate the specificity of the type III pentaketide synthase from Neurospora crassa.
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DOI：
10.1016/j.chembiol.2008.08.011
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发表时间：
2008-10-20
影响因子：
--
通讯作者：
Nair SK
中科院分区：
文献类型：
--
作者：
Rubin-Pitel SB;Zhang H;Vu T;Brunzelle JS;Zhao H;Nair SK
III 型聚酮合酶 (PKS) 查尔酮合酶超家族中的酶可产生多种聚酮化合物，这些化合物在结构和生物功能方面各不相同。第一个被表征的真菌 III 型 PKS 是粗糙脉孢菌 2'-氧代烷基间苯二甲酸合酶 (ORAS)，它被证明可以用一系列酰基辅酶 A 硫酯 (C4-C20) 引发，并使用丙二酰辅酶 A 延伸，从而产生吡喃酮、间苯二酚和间苯二酚产品。为了进一步了解这种不寻常的底物特异性和独特的产物特征，我们以 1.75 Å 分辨率测定了催化活性截短 ORAS 的晶体结构，以 2.1 Å 分辨率测定了 Phe-252→Gly 定点突变体，以及以 2.0 Å 分辨率测定了 ORAS 与二十烷酸的二元复合物的晶体结构。晶体结构揭示了酶活性位点附近结构元件的明显重排，允许容纳长链脂肪酸酯，并重新定向用于控制环化和聚酮化合物链长度的门控机制。通过许多基于结构的定点变体的功能表征进一步阐明了这些新颖结构元件的作用。这些研究确立了 PKS 折叠的先前意想不到的可塑性，这是对该酶家族其他成员的结构研究所没有预料到的。这些架构元素为旨在利用 III 型 PKS 多样性的进一步工程实验提供了额外的目标。
Enzymes in the chalcone synthase superfamily of type III polyketide synthases (PKSs) produce a wide variety of polyketide compounds, distinct in terms of both structure and biological function. The first fungal type III PKS to be characterized, Neurospora crassa 2′-oxoalkylresorcylic acid synthase (ORAS), was demonstrated to prime with a range of acyl-Coenzyme A thioesters (C4—C20) and extend using malonyl-Coenzyme A, thereby producing pyrone, resorcinol, and resorcylic acid products. To gain further insight into this unusual substrate specificity and unique product profile, we have determined the crystal structures of a catalytically active truncated ORAS to 1.75 Å resolution, of the Phe-252→Gly site-directed mutant to 2.1 Å resolution, and of a binary complex of ORAS with eicosanoic acid to 2.0 Å resolution. The crystal structures reveal a distinct rearrangement of structural elements near the enzyme active site that allows for accommodation of long chain fatty acid esters and a reorientation of the gating mechanism for controlling cyclization and polyketide chain length. The roles of these novel structural elements are further elucidated by functional characterization of a number of structure-based site-directed variants. These studies establish a previously unexpected plasticity to the PKS fold, unanticipated from structural studies of other members of this enzyme family. These architectural elements provide an additional target for further engineering experiments aimed at exploiting the diversity of type III PKSs.
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DOI：
10.1038/11472
发表时间：
1999-08-01
期刊：
NATURE STRUCTURAL BIOLOGY
影响因子：
--
作者：
Schröder, J
通讯作者：
Schröder, J
DOI：
10.1016/j.bbrc.2005.03.160
发表时间：
2005-05-27
影响因子：
3.1
作者：
Seshime, Y;Juvvadi, PR;Kitamoto, K
通讯作者：
Kitamoto, K
DOI：
10.1107/s0907444903018043
发表时间：
2003-11-01
期刊：
ACTA CRYSTALLOGRAPHICA SECTION D-BIOLOGICAL CRYSTALLOGRAPHY
影响因子：
--
作者：
Grosse-Kunstleve, RW;Adams, PD
通讯作者：
Adams, PD
影响因子：
15
作者：
Abe, I;Utsumi, Y;Noguchi, H
通讯作者：
Noguchi, H
影响因子：
5.6
作者：
VANDUYNE, GD;STANDAERT, RF;CLARDY, J
通讯作者：
CLARDY, J