Silvaroli Josie A, Pleshinger Matthew J, Banerjee Surajit, Kiser Philip D, Golczak Marcin
Department of Pharmacology, School of Medicine, Case Western Reserve University , Cleveland, Ohio, United States.
College of Wooster , Wooster, Ohio, United States.
ACS Chem Biol. 2017 Sep 15;12(9):2296-2304. doi: 10.1021/acschembio.7b00336. Epub 2017 Jul 26.
The biochemical pathway that gives onions their savor is part of the chemical warfare against microbes and animals. This defense mechanism involves formation of a volatile lachrymatory factor (LF) ((Z)-propanethial S-oxide) that causes familiar eye irritation associated with onion chopping. LF is produced in a reaction catalyzed by lachrymatory factor synthase (LFS). The principles by which LFS facilitates conversion of a sulfenic acid substrate into LF have been difficult to experimentally examine owing to the inherent substrate reactivity and lability of LF. To shed light on the mechanism of LF production in the onion, we solved crystal structures of LFS in an apo-form and in complex with a substrate analogue, crotyl alcohol. The enzyme closely resembles the helix-grip fold characteristic for plant representatives of the START (star-related lipid transfer) domain-containing protein superfamily. By comparing the structures of LFS to that of the abscisic acid receptor, PYL10, a representative of the START protein superfamily, we elucidated structural adaptations underlying the catalytic activity of LFS. We also delineated the architecture of the active site, and based on the orientation of the ligand, we propose a mechanism of catalysis that involves sequential proton transfer accompanied by formation of a carbanion intermediate. These findings reconcile chemical and biochemical information regarding thioaldehyde S-oxide formation and close a long-lasting gap in understanding of the mechanism responsible for LF production in the onion.
赋予洋葱风味的生化途径是其对抗微生物和动物的化学防御机制的一部分。这种防御机制涉及一种挥发性催泪因子(LF)((Z)-丙烷硫醛S-氧化物)的形成,它会导致切洋葱时常见的眼睛刺激。LF是在催泪因子合酶(LFS)催化的反应中产生的。由于LF固有的底物反应性和不稳定性,LFS促进亚磺酸底物转化为LF的原理一直难以通过实验进行研究。为了阐明洋葱中LF产生的机制,我们解析了LFS的无配体形式以及与底物类似物巴豆醇复合物的晶体结构。该酶与含START(与星状相关的脂质转移)结构域的蛋白质超家族中植物代表所特有的螺旋握把折叠结构非常相似。通过将LFS的结构与START蛋白质超家族的代表——脱落酸受体PYL10的结构进行比较,我们阐明了LFS催化活性背后的结构适应性。我们还描绘了活性位点的结构,并根据配体的方向,提出了一种催化机制,该机制涉及伴随碳负离子中间体形成的顺序质子转移。这些发现整合了有关硫代醛S-氧化物形成的化学和生化信息,填补了长期以来在理解洋葱中LF产生机制方面的空白。