Department of Medicinal Chemistry, University of Washington, Seattle, WA 98195-7610, USA.
Biomolecules. 2023 Feb 9;13(2):329. doi: 10.3390/biom13020329.
High concentrations of electrophilic lipid alkenals formed during oxidative stress are implicated in cytotoxicity and disease. However, low concentrations of alkenals are required to induce antioxidative stress responses. An established clearance pathway for lipid alkenals includes conjugation to glutathione (GSH) via Michael addition, which is catalyzed mainly by glutathione transferase isoform A4 (GSTA4-4). Based on the ability of GSTs to catalyze hydrolysis or -Michael addition of GSH conjugates, and the antioxidant function of low concentrations of lipid alkenals, we hypothesize that GSTA4-4 contributes a homeostatic role in lipid metabolism. Enzymatic kinetic parameters for -Michael addition with trans-2-Nonenal (NE) reveal the chemical competence of GSTA4-4 in this putative role. The forward GSTA4-4-catalyzed Michael addition occurs with the rapid exchange of the C2 proton of NE in DO as observed by NMR. The isotope exchange was completely dependent on the presence of GSH. The overall commitment to catalysis, or the ratio of first order k for 'forward' Michael addition to the first order k for H/D exchange is remarkably low, approximately 3:1. This behavior is consistent with the possibility that GSTA4-4 is a regulatory enzyme that contributes to steady-state levels of lipid alkenals, rather than a strict 'one way' detoxication enzyme.
氧化应激过程中形成的高浓度亲电性脂质烯醛与细胞毒性和疾病有关。然而，诱导抗氧化应激反应需要低浓度的烯醛。脂质烯醛的一种已建立的清除途径包括通过迈克尔加成与谷胱甘肽 (GSH) 缀合，主要由谷胱甘肽转移酶同工酶 A4 (GSTA4-4) 催化。基于 GST 催化 GSH 缀合物水解或 -迈克尔加成的能力，以及低浓度脂质烯醛的抗氧化功能，我们假设 GSTA4-4 在脂质代谢中发挥着动态平衡作用。用反式-2-壬烯醛 (NE) 进行 -迈克尔加成的酶动力学参数揭示了 GSTA4-4 在这种假定作用中的化学能力。通过 NMR 观察到，正向 GSTA4-4 催化的迈克尔加成伴随着 NE 的 C2 质子在 DO 中的快速交换。同位素交换完全依赖于 GSH 的存在。总体催化承诺，或“正向”迈克尔加成的一级 k 与 H/D 交换的一级 k 的比值非常低，约为 3:1。这种行为与 GSTA4-4 可能是一种调节酶的可能性一致，它有助于脂质烯醛的稳态水平，而不是严格的“单向”解毒酶。
