二叶酸减少酶如何促进二叶酸的质子化
Thomas H Rod1, Charles L Brooks
1Department of Molecular Biology, TPC6, 10550 North Torrey Pines Road, La Jolla, California 92037, USA.
Journal of the American Chemical Society
|July 17, 2003
概括
双叶酸还原酶 (DHFR) 促进了双叶酸 (H2F) 的4 pKa单位的质子化,增强了它的活性. 这通过涉及Asp27和疏水活性位点的静电相互作用发生.
科学领域:
- 生物化学 生物化学
- 酶动力学 酶动力学
- 计算化学是一种计算化学.
背景情况:
- 双叶酸还原酶 (DHFR) 对于叶酸代谢至关重要,催化二叶酸 (H2F) 减少为四叶酸.
- DHFR活动的pH依赖性表明,质子化事件在催化机制中起着作用.
- 了解这些机制对于开发针对DHFR的药物至关重要.
研究的目的:
- 在DHFR活性位点内确定N5原子在H2F中的pKa值.
- 阐明负责DHFR促进H2F质子化的分子机制.
- 解释活性部位残留物和酶构成对催化效率的影响.
主要方法:
- 封闭和封闭形状的DHFR/NADPH/H2F复合物的分子动力学 (MD) 模拟 (10-12.5 ns).
- 自由能量扰动 (FEP) 计算以确定H2F中的N5原子的pKa.
- 分析静电相互作用和活性部位的疏水性.
主要成果:
- 在DHFR催化反应中,H2F的N5原子被确定为pH依赖的关键位置.
- DHFR可使H2F的质子化增加约4 pKa单位.
- 在Asp27和N5的质子之间有利的静电相互作用,加上疏水性M20循环的增强,推动了这种效应.
- 酶诱导的H2F构造也有利于N5质子化.
结论:
- DHFR积极促进H2F的质子化,解释其pH取决于pH的活性.
- Asp27残留物和疏水活性部位在增强质子化方面发挥着关键作用.
- 这些发现支持了先前的研究,并解释了在中性pH下Asp27突变的活性降低.
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