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结构和鉴定来自Methanosarcina acetivorans lysate的原生PLP合成复合物的结构和鉴定

Angela Agnew^1,2,3, Ethan Humm¹, Kang Zhou²

¹Department of Microbiology, Immunology & Molecular Genetics, University of California Los Angeles, Los Angeles, California, USA.

|November 26, 2024

概括

我们使用cryo-EM.确定了来自古生物的pyridoxal 5'-phosphate (PLP) 合成酶的原生结构. 这种方法揭示了原生蛋白质状态,克服了研究这些关键生物体的挑战.

科学领域:

结构生物学结构生物学
生物化学生化学
微生物学微生物学

背景情况:

蛋白质结构可以在纯化形式和它们的体内状态之间显著不同.
甲原体古生物对全球碳循环和人类微生物群至关重要,但它们的酶结构的特征很差.
了解原生考古蛋白质结构对于阐明它们的功能和影响至关重要.

研究的目的:

应用cryoID方法来确定甲原性古物*Methanosarcina acetivorans*中的pyridoxal 5 -phosphate (PLP) 合成酶 (PdxS) 的原生结构.
研究PdxS的原生四级结构,并将其与之前确定的结构进行比较.
为了证明cryoID的实用性,以捕捉考古系统中的原生结构状态.

主要方法:

应用了自下而上的结构蛋白质组学方法,cryoID.
使用低温电子显微镜 (cryo-EM) 来分析来自*Methanosarcina acetivorans*的分离细胞溶解物.
从冷EM数据中重建和识别本地PdxS复合物.

主要成果:

本地PdxS复合体成功地被重建成一个homo-dodecamer.
在细胞溶解物中没有观察到PdxS和PdxT的先前提出的超复合体.
原生PdxS单体结构揭示了具有额外螺旋体的8α/8β TIM-桶域,因缺乏37-氨基酸插入而与先前的晶体结构不同. 在活性部位发现了ribose-5-phosphate.

结论:

化ID工作流程使得从异质细胞样本中以原子分辨率确定原生蛋白质结构.
*M. acetivorans* PdxS的原生结构不同于之前报告的结构,突出了体内结构确定的重要性.
这项研究推进了考古酶的结构特征,这对于理解它们在环境和健康环境中的作用至关重要.

关键词:

甲沙辛酸乙基甲沙辛酸乙基甲沙辛酸乙基在PLP合成酶中.低温电磁波冷却器 (Cryo-EM) 是一个非常好的方法.这种冷化器是 cryoID 的.

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