MEASUREMENT OF THE ISOTOPE ENRICHMENT OF STABLE ISOTOPE-LABELED PROTEINS USING

这个子项目是利用由NIH/NCRR资助的中心拨款提供的资源的许多研究子项目之一。子项目和调查员(PI)可能从另一个NIH来源获得了主要资金，因此可能会出现在其他CRISE条目中。列出的机构是针对中心的，而不一定是针对调查员的机构。稳定的同位素富集物分子被用作体内标准物质和体内底物代谢的示踪剂。质谱仪中测量的比例与摩尔比例的准确转换是复杂的，因为含有浓缩原子的多原子分子将导致同位素异构体的组合分布，这取决于“标记的”原子的浓缩程度和数量。这种影响可能会在摩尔比测量中造成很大的误差，这取决于使用哪个或哪些同位素峰来确定比率。我们报道了一种计算方法，它预测了一定范围内的同位素分布，并将预测的分布与傅里叶变换离子回旋共振质谱仪获得的多肽同位素分布进行了比较。我们的方法是准确的，测量的丰度在预期同位素分布的1.5%以内。对于原子%超过15N的59、79和99个多肽，该方法的相对标准偏差分别为4.9、2.0和0.8%。这种方法是自动化的，使得在一次LC-FTICR-MS实验中对数千个多肽进行同位素浓缩计算成为可能。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Stable isotope-enriched molecules are used as internal standards and as tracers of in vivo substrate metabolism. The accurate conversion of measured ratios in the mass spectrometer to mole ratios is complicated because a polyatomic molecule containing enriched atoms will result in a combinatorial distribution of isotopomers depending on the enrichment and number of "labeled" atoms. This effect could potentially cause a large error in the mole ratio measurement depending on which isotope peak or peaks were used to determine the ratio. We report a computational method that predicts isotope distributions over a range of enrichments and compares the predicted distributions to experimental peptide isotope distributions obtained by Fourier transform ion cyclotron resonance mass spectrometry. Our approach is accurate with measured enrichments within 1.5% of expected isotope distributions. The method is also precise with 4.9, 2.0, and 0.8% relative standard deviations for peptides containing 59, 79, and 99 atom % excess 15N, respectively. The approach is automated making isotope enrichment calculations possible for thousands of peptides in a single LC-FTICR-MS experiment.