Jagiellonian University, Ingardena, Krakow, Poland.
Bioorg Chem. 2010 Jun;38(3):132-7. doi: 10.1016/j.bioorg.2010.02.002. Epub 2010 Feb 13.
The aim of this study was elucidation of the difference in inhibition influence of 5-hydroxy-1,4-naphthoquinone (juglone) and 2-hydroxy-1,4-naphthoquinone (lawsone) on jack bean urease activity. It was found that juglone acted as a strong, time and concentration dependent inactivator of urease. On the contrary, lawsone showed an inconsiderable inhibition influence. The reactivation of juglone modified urease showed the participation of reversible and irreversible contribution in the inactivation. In the presence of an excess of DTT, urease inactivated by juglone regained 70% of its activity. The reversible inactivation was attributed to oxidation of the essential urease thiols by reactive oxygen species (ROS) realizing during reduction of juglone to seminaphthoquinone. Presence of hydrogen peroxide in the incubation system was proved by direct determination and by application of catalase. The irreversible contribution in the inhibition was assumed as an arylation of urease thiol groups by juglone. The insignificant urease inhibition by lawsone was concluded as an effect of a low hydrogen peroxide generation and lawsone resistance for reaction with protein thiols. It was found that lawsone well reacted with l-cysteine, poorly with glutathione and hardly with urease thiols. The observed sequence was arranged according the rule the more complex thiol the less susceptible for reaction with lawsone. On the other hand, juglone displayed an excellent reactivity towards both thiols and urease. Thus, this indicated a significance of a steric hindrance which appeared when the hydroxyl group changing position from 5 in juglone (5-hydroxy-1,4-naphthoquinone) to 2 in lawsone (2-hydroxy-1,4-naphthoquinone).
本研究旨在阐明 5-羟基-1,4-萘醌（胡桃醌）和 2-羟基-1,4-萘醌（洛索纳尔）对菜豆脲酶活性抑制影响的差异。结果发现，胡桃醌是一种强的、时间和浓度依赖性的脲酶抑制剂。相比之下，洛索纳尔的抑制作用则可以忽略不计。胡桃醌修饰脲酶的复活动力学表明，其失活既有可逆的也有不可逆的贡献。在过量 DTT 的存在下，被胡桃醌失活的脲酶恢复了其 70%的活性。可逆失活归因于在还原为半萘醌的过程中，活性氧（ROS）氧化脲酶必需的巯基。孵育体系中过氧化氢的存在通过直接测定和应用过氧化氢酶得到证实。不可逆失活被认为是胡桃醌对脲酶巯基的芳基化。洛索纳尔对脲酶的抑制作用不明显，这被认为是由于其产生的过氧化氢量低，以及洛索纳尔对蛋白质巯基的反应性低所致。结果发现，洛索纳尔与 l-半胱氨酸反应良好，与谷胱甘肽反应较差，与脲酶巯基反应几乎没有。观察到的序列是根据规则排列的，即较复杂的巯基对洛索纳尔的反应性越低。另一方面，胡桃醌对两种硫醇和脲酶都表现出良好的反应性。因此，这表明当羟基从胡桃醌的 5 位（5-羟基-1,4-萘醌）变为洛索纳尔的 2 位（2-羟基-1,4-萘醌）时，空间位阻的出现具有重要意义。
