Institute of Biochemistry and Molecular Biology, National Yang-Ming University, Taipei 112, Taiwan.
Anal Biochem. 2013 Sep 1;440(1):108-13. doi: 10.1016/j.ab.2013.05.018. Epub 2013 Jun 4.
While recent developments in mass spectrometry enable direct evaluation of monoisotopic masses (M(mi)) of smaller compounds, protein M(mi) is mostly determined based on its relationship to average mass (Mav). Here, we propose an alternative approach to determining protein M(mi) based on its correlation with the most abundant mass (M(ma)) measurable using high-resolution mass spectrometry. To test this supposition, we first empirically calculated M(mi) and M(ma) of 6158 Escherichia coli proteins, which helped serendipitously uncover a linear correlation between these two protein masses. With the relationship characterized, liquid chromatography-mass spectrometry was employed to measure M(ma) of protein samples in its ion cluster with the highest signal in the mass spectrum. Generally, our method produces a short series of likely M(mi) in 1-Da steps, and the probability of each likely M(mi) is assigned statistically. It is remarkable that the mass error of this M(mi) is as miniscule as a few parts per million, indicating that our method is capable of determining protein M(mi) with high accuracy. Benefitting from the outstanding performance of modern mass spectrometry, our approach is a significant improvement over others and should be of great utility in the rapid assessment of protein primary structures.
虽然质谱分析的最新进展可以直接评估较小化合物的单一同位素质量 (M(mi))，但蛋白质的 M(mi) 主要基于其与平均质量 (Mav) 的关系来确定。在这里，我们提出了一种替代方法，基于其与使用高分辨率质谱可测量的最丰富质量 (M(ma)) 的相关性来确定蛋白质的 M(mi)。为了验证这一假设，我们首先通过经验计算了 6158 种大肠杆菌蛋白质的 M(mi)和 M(ma)，这偶然揭示了这两种蛋白质质量之间存在线性相关性。在确定了这种关系后，我们使用液相色谱-质谱法测量了在质谱中信号最强的蛋白质离子簇中的 M(ma)。通常，我们的方法以 1-Da 的步长产生一系列可能的 M(mi)，并且每个可能的 M(mi) 的概率都是通过统计学分配的。值得注意的是，这种 M(mi) 的质量误差小到百万分之几，这表明我们的方法能够非常准确地确定蛋白质的 M(mi)。得益于现代质谱分析的卓越性能，我们的方法比其他方法有了显著的改进，应该在快速评估蛋白质一级结构方面具有很大的实用性。
