碳酸盐合成酶中的碳酸盐运输:一个理论和实验研究
Liliya Lund1, Yubo Fan, Qiang Shao
1Department of Chemistry, Texas A&M University, College Station, Texas 77843-3255, USA.
Journal of the American Chemical Society
|March 2, 2010
概括
碳酸合成酶 (CPS) 使用道在酶内运输碳酸盐. 分子动力学和突变发生揭示了对于这种运输至关重要的能量障碍和水口袋,突变显著降低了酶活性.
科学领域:
- 生物化学 生物化学
- 结构生物学 结构生物学
- 酶催化酶的催化作用
背景情况:
- 碳酸合成酶 (CPS) 催化了核酸和氨基酸生物合成的关键步骤.
- 碳酸盐是一种中间体,必须在酶的大子单元内从其形成地点到利用地点有效地运输.
- 了解这种传输机制对于阐明酶的功能和调节至关重要.
研究的目的:
- 通过大肠杆菌CPS的大小单元研究碳酸盐的运输途径.
- 确定控制碳酸盐转移的关键结构特征和能源景观.
- 通过突变生成验证特定道区域在酶活性中的作用.
主要方法:
- 用分子动力学 (MD) 模拟来建模碳酸盐运输.
- 平均力 (PMF) 计算的潜力被用来确定沿运输道的自由能量配置文件.
- 在拟议的碳酸盐道内对关键残留物进行了局部定向的突变发生.
主要成果:
- 碳酸盐运输道由三个水口袋和两个狭窄的狭窄组成,形成6.7和8.4kcal/mol的自由能量障碍.
- 碳酸盐转位是由包含Arg-306在酸盐释放时旋转的形状变化促进的,打开道.
- 设计用于阻塞狭窄道部分的A23F和G575F突变体,具有显著降低的催化活性 (分别为野生类型的1.7%和3.8%).
结论:
- 大肠杆菌CPS中的碳酸盐道存在重要的能量障碍,影响转移效率.
- 形状的灵活性,特别是Arg-306的旋转,对于打开道和促进碳酸盐通道至关重要.
- 道最狭窄的部分内的特定残留物对于维持CPS的催化活性至关重要.
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