拉尔B基质复合物的结构和在核酸辅因子生物合成过程中识别反应中间体
Shramana Chatterjee1, Jorge L Nevarez2, Joel A Rankin1
1Department of Microbiology and Molecular Genetics, Michigan State University, East Lansing, Michigan 48824, United States.
Biochemistry
|October 13, 2023
概括
大B酶使用cysteinyl链接到NAD+用于-钉辅因子生物合成. 新的结构和光谱数据显示了一种经过修订的催化机制,涉及CO2固定和基质碳素化.
科学领域:
- 生物化学 生物化学
- 酶学 是一种酶学.
- 结构生物学 结构生物学
背景情况:
- 拉尔B对于-子辅助因子生物合成至关重要,催化尼古丁酸腺因二核酸 (NaAD) 转化为AMP和-3,5-双碳酸单核酸 (P2CMN).
- 之前的研究表明,LarB利用二氧化碳进行炭化,并揭示了Lactiplantibacillus plantarum LarB中Cys221和NAD+之间的共价链接.
- 这种联系被假设为促进C5碳素化,激活水用于酸水解.
研究的目的:
- 为了进一步证明野生类型LarB中的cysteinyl与NAD+的联系,确认它不是一个结晶工件.
- 研究S127A LarB变体与NaAD的相互作用,并确定反应中间体.
- 通过结构和光谱分析阐明LarB的催化机制.
主要方法:
- 紫外可见光谱检测氨酸侧链与NAD+的连接.
- 质谱测量以确定反应中间体,特别是丁尼科丁酸腺因二核酸 (DaAD).
- 进行X射线晶体学以确定S127A LarB·NaAD复合物的结构.
主要成果:
- 光谱数据证实了野生型Larb.B.中的cysteinyl与NAD+的联系.
- 通过使用S127A变种识别了中间的迪尼科丁酸腺因二核酸 (DaAD).
- S127A LarB·NaAD 的晶体结构提供了 CO2 结合的洞察力,并揭示了 Glu180 的定位,表明它不参与 C5 质子抽象.
结论:
- 囊与NAD+的链接是LarB活动的真正特征.
- 该S127A变种促进了关键中间体的识别,并为机制提供了结构基础.
- 根据综合的结构,光谱和生物化学数据,建议对LarB进行修订的催化机制.
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