格拉米西丁S合成酶的启动模块PheATE的基质识别和选择
L Luo1, M D Burkart, T Stachelhaus
1Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, 240 Longwood Avenue, Boston, Massachusetts 02115, USA.
Journal of the American Chemical Society
|November 8, 2001
概括
非核糖体合成酶的PheATE启动模块激活L-氨酸和其他氨基酸. 表皮化域作为一个守门员,控制D-phenylalanine的形成,用于gramicidin S的合成.
科学领域:
- 生物化学 生物化学
- 分子生物学分子生物学
- 酶学 是一种酶学.
背景情况:
- 非核糖体合成酶 (NRPS) 是大型的酶复合体,可以合成各种天然产品的.
- NRPS的启动模块对于选择和激活第一个氨基酸至关重要,从而启动组装线.
- 格拉米西丁S合成酶启动模块 (PheATE) 激活L-氨 (L-Phe) 并开始合成抗生素格拉米西丁S.
研究的目的:
- 为了研究PheATE腺化域超出其主要基质L-Phe.的基质选择性.
- 了解PheATE启动模块对基质识别和选择的动力学和热力学基础.
- 阐明表皮化域在控制D-aminoacyl中间体的形成中的作用.
主要方法:
- 测试各种蛋白质生成氨基酸的基化域的选择性.
- 进行单次转换的动力学研究,以确定微观反应速率和平衡常量.
- 构建自由能量配置文件以分析基质识别和反应机制.
主要成果:
- 除了L-Phe之外,PheATE腺化域还激活了L-氨酸 (L-Tyr),L-三氨酸 (L-Trp) 和L-氨酸 (L-Leu).
- 没有观察到非同源氨基酸-腺-5'-单酸盐 (氨基酸-AMP) 或氨基酸-酶中间体的水解编辑.
- 与L-Phe.相比,L-Trp,L-Tyr和L-Leu对它们的D-enantiomers的表皮化率显著降低 (245至540倍).
结论:
- PheATE 腺化域表现出比以前假设的更广泛的基质特异性.
- 表皮化域充当关键的守门员,确保有效生成所需的D-aminoacyl-酶,用于gramicidin S生物合成.
- 动力学和热力学分析提供了对NRPS基质选择的分子机制的洞察.
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