Sensitive determination of metalloprotein in salt-rich matrices by size exclusion chromatography coupled with inductively coupled plasma-mass spectrometry
Sensitive determination of metalloprotein in salt-rich matrices by size exclusion chromatography coupled with inductively coupled plasma-mass spectrometry
复制标题
DOI：
10.1016/j.chroma.2022.463303
复制
发表时间：
2022-07-10
影响因子：
4.1
通讯作者：
Jiang, Guibin
中科院分区：
文献类型：
--
作者：
Tang, Yinyin;Liu, Lihong;Jiang, Guibin
金属蛋白质在各种严重依赖金属离子和各种蛋白质的生物过程中起着至关重要的作用。然而，目前仍缺乏快速分析复杂样品中金属蛋白的方法，特别是在富盐基质中。本研究基于体积排阻色谱-电感耦合等离子体质谱联用技术（SEC-ICP-MS），结合生物体系中必不可少的高基质引入（HMI）模式，建立了一种高灵敏度的富盐基质中金属蛋白的分离测定方法。以牛血清白蛋白（BSA，66.0 kDa）、卵清蛋白（OVA，44.0 kDa）、碳酸酐酶（CA，29.0 kDa）和核糖核酸酶A（RA，13.7 kDa）4种不同分子量的碘标记蛋白质，优化了SEC-ICP-MS系统的分离条件。优化后，以10 mmol/L HEPES和40 mmol/L Na 2SO 4（pH=7.0）为移动的相，在30 min内成功分离了4种碘标记蛋白质和碘离子，并建立了分子量log与保留时间的线性关系。4种碘标记蛋白质的保留时间和峰面积的相对标准偏差（RSD，n = 5）分别在0.2-0.9%和3.3- 7.7%之间，表明精密度和重复性良好。将该方法成功应用于真实的生物组织样品中铅结合蛋白的快速分离和检测。(C)2022由爱思唯尔公司出版
Metalloproteins play crucial and distinct roles in a variety of biological processes that rely heavily on the metal ions and various proteins. However, there is still a lack of method for rapid analysis of metalloproteins in complex samples, especially in salt-rich matrices. In this study, a sensitive method for separation and determination of metalloproteins in salt-rich matrices was developed based on the size exclusion chromatography coupled with inductively coupled plasma-mass spectrometry (SEC-ICP-MS), combining with the high matrix introduction (HMI) mode, which is quite essential for biological system. The separation conditions of the SEC-ICP-MS system were optimized by using four iodine labeled proteins with different molecular weights, including bovine serum albumin (BSA, 66.0 kDa), ovalbumin (OVA, 44.0 kDa), carbonic anhydrase (CA, 29.0 kDa) and ribonuclease A (RA, 13.7 kDa). After optimization, four iodine labeled proteins and iodine ions were successfully separated within 30 min by using 10 mmol/L HEPES and 40 mmol/L Na2SO4 (pH=7.0) as mobile phase and a linear relationship between log molecular weight and retention time was established. The relative standard deviations (RSDs, n = 5) of the retention time and peak areas for the four iodine labeled proteins were in the range of 0.2-0.9% and 3.3-7.7%, respectively, suggesting good precision and repeatability. Then the proposed method was successfully applied to the rapid separation and detection of lead-binding proteins in real biological tissue samples. (C) 2022 Published by Elsevier B.V.