通过质子核磁共振光谱学对可溶性蛋白质的绝对定量:血中蛋白质总度
Panteleimon G Takis1,2,3, Ivan Vučković4, Anna M Kowalka5
1Section of Bioanalytical Chemistry, Division of Systems Medicine, Department of Metabolism, Digestion, and Reproduction, Imperial College London, London SW7 2AZ, U.K.
Analytical chemistry
|October 4, 2024
概括
核磁共振 (NMR) 光谱可以通过将它们视为同聚合物来量化蛋白质. 这种方法在没有校准标准的情况下为蛋白质定量化建立了通用转换常数,利用NMR光谱的甲基区域.
科学领域:
- 生物化学 生物化学
- 分析化学 分析化学
- 频谱学是一种光谱学.
背景情况:
- 绝对蛋白质的量化在各种科学领域至关重要.
- 像基尔达尔分析这样的当前方法有局限性.
- 核磁共振 (NMR) 谱学为蛋白质分析提供了潜力.
研究的目的:
- 开发一种使用NMR光谱学的绝对蛋白质量定量方法.
- 建立用于蛋白质量化的通用转换常量.
- 用合成光谱和真实世界样本 (如血) 来验证该方法.
主要方法:
- 蛋白质被建模为具有有效氨基酸残留物的同聚合物.
- 确定了每个残留物的分子量和含量.
- 1H NMR光谱被合成并与实验数据进行比较.
- 对各种样本计算了甲基区域的实证比率.
主要成果:
- 在有效氨基酸残留物质之间发现了一个常数比率 (14.3 ± 0.3),类似于Kjeldahl常数.
- 合成的蛋白质光谱显示,甲基区域的蛋白质质量与质子比为1 (mg/mL) /mM.
- 在人血中进行的实验验证,得到了一致的比率 (1.115 ± 0.006 (mg/mL) /mM).
结论:
- 一个1H的NMR频谱的总积分可以用于总蛋白质的量化.
- 通过NMR量化蛋白质存在通用转换常量,独立于蛋白质组成.
- 这种基于NMR的方法可以在没有校准标准的情况下准确量化蛋白质.
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