一个保存的第二个球体残留在Lytic多糖单氧酶中的铜位反应性
Kelsi R Hall1, Chris Joseph2, Iván Ayuso-Fernández1
1Faculty of Chemistry, Biotechnology and Food Science, Norwegian University of Life Sciences (NMBU), 1432, Ås, Norway.
Journal of the American Chemical Society
|August 16, 2023
概括
在铜中心附近保存的谷氨酸/谷氨酸酸残留微调了酸多糖单氧酶 (LPMO) 功能. 突变影响酶的反应性,稳定性和催化机制,为LPMO活动和合成催化剂设计提供了洞察力.
科学领域:
- 生物化学
- 酵素学
- 生物有机化学
背景情况:
- 酸多糖单氧酶 (LPMOs) 是一种铜酶,其激活C-H键的机制尚不清楚.
- 一个保存的谷氨酸/谷氨酸残留物位于铜中心的第二个协调球.
研究的目的:
- 研究保存的谷氨酸/谷氨酸残留在LPMO功能和铜反应性的作用.
- 确定该残留物中的突变如何影响LPMO的催化机制和稳定性.
主要方法:
- 在*Nc*AA9C中的Gln残留物转化为Glu,Asp或Asn.
- 酶活性测定,包括降解潜力和再氧化率测量.
- 电子磁共振 (EPR) 和X射线吸收光谱 (XAS).
- 密度函数理论 (DFT) 的计算.
主要成果:
- 突变改变了LPMO的功能和铜的反应性,Glu/Asp的接近降低了降解潜力和降低了降解/再氧化速率的比率.
- 突变者表现出增加的酶失活和改变的保护性跳洞通路.
- EPR和XAS显示突变没有直接扰乱Cu (II) 配体场.
- DFT的计算表明,Glu的质子化增强了再氧化速率,并揭示了质子化Glu形式的Cu (III) - 氧化物中间体.
结论:
- 第二个球的谷氨酸/谷氨酸残留物是LPMO催化功能的关键决定因素.
- 这种残留物影响铜的反应性,酶的稳定性和反应中间体.
- 这些发现为了解LPMOs和设计新型合成催化剂提供了洞察力.
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