Koglin Alexander, Löhr Frank, Bernhard Frank, Rogov Vladimir V, Frueh Dominique P, Strieter Eric R, Mofid Mohammad R, Güntert Peter, Wagner Gerhard, Walsh Christopher T, Marahiel Mohamed A, Dötsch Volker
Institute of Biophysical Chemistry and Center for Biomolecular Magnetic Resonance and Cluster of Excellence Macromolecular Complexes (CEF), J.W.-Goethe University, 60438 Frankfurt am Main, Germany.
Nature. 2008 Aug 14;454(7206):907-11. doi: 10.1038/nature07161.
Non-ribosomal peptide synthetases (NRPS) and polyketide synthases (PKS) found in bacteria, fungi and plants use two different types of thioesterases for the production of highly active biological compounds. Type I thioesterases (TEI) catalyse the release step from the assembly line of the final product where it is transported from one reaction centre to the next as a thioester linked to a 4'-phosphopantetheine (4'-PP) cofactor that is covalently attached to thiolation (T) domains. The second enzyme involved in the synthesis of these secondary metabolites, the type II thioesterase (TEII), is a crucial repair enzyme for the regeneration of functional 4'-PP cofactors of holo-T domains of NRPS and PKS systems. Mispriming of 4'-PP cofactors by acetyl- and short-chain acyl-residues interrupts the biosynthetic system. This repair reaction is very important, because roughly 80% of CoA, the precursor of the 4'-PP cofactor, is acetylated in bacteria. Here we report the three-dimensional structure of a type II thioesterase from Bacillus subtilis free and in complex with a T domain. Comparison with structures of TEI enzymes shows the basis for substrate selectivity and the different modes of interaction of TEII and TEI enzymes with T domains. Furthermore, we show that the TEII enzyme exists in several conformations of which only one is selected on interaction with its native substrate, a modified holo-T domain.
在细菌、真菌和植物中发现的非核糖体肽合成酶（NRPS）和聚酮化合物合成酶（PKS）利用两种不同类型的硫酯酶来生产高活性生物化合物。I型硫酯酶（TEI）催化最终产物从装配线上释放，在装配线上，最终产物作为与共价连接到硫醇化（T）结构域的4'-磷酸泛酰巯基乙胺（4'-PP）辅因子相连的硫酯从一个反应中心转运到下一个反应中心。参与这些次生代谢物合成的第二种酶，即II型硫酯酶（TEII），是NRPS和PKS系统全T结构域功能性4'-PP辅因子再生的关键修复酶。4'-PP辅因子被乙酰基和短链酰基残基错误引发会中断生物合成系统。这种修复反应非常重要，因为在细菌中，4'-PP辅因子的前体辅酶A大约80%被乙酰化。在此，我们报道了来自枯草芽孢杆菌的II型硫酯酶的三维结构，该结构处于游离状态以及与一个T结构域结合的状态。与TEI酶结构的比较显示了底物选择性的基础以及TEII和TEI酶与T结构域相互作用的不同模式。此外，我们表明TEII酶以几种构象存在，其中只有一种构象在与它的天然底物（一种修饰的全T结构域）相互作用时被选择。
