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

使用测量稳定同位素标记蛋白质的同位素富集

• 批准号: 7420823
• 负责人: John R Yates III
• 金额: $ 3.67万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-09-20 至 2007-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7420823
• 关键词: ions; mass spectrometry; measurement; metabolism; peptides; stable isotope

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.

本子项目是利用由NIH/NCRR资助的中心赠款提供的资源的众多研究子项目之一。子项目和研究者（PI）可能已经从另一个NIH来源获得了主要资金，因此可以在其他CRISP条目中表示。列出的机构是中心的，不一定是研究者的机构。稳定的同位素富集分子被用作体内底物代谢的内标和示踪剂。质谱仪中测量的比值与摩尔比的精确转换是复杂的，因为含有富集原子的多原子分子将导致同位素体的组合分布，这取决于“标记”原子的富集程度和数量。这种效应可能会在测量摩尔比时造成很大的误差，这取决于使用哪个同位素峰或哪个峰来确定摩尔比。我们报告了一种计算方法，该方法预测了在一定范围内富集的同位素分布，并将预测分布与傅立叶变换离子回旋共振质谱法获得的实验肽同位素分布进行了比较。我们的方法是准确的，测量的富集在预期同位素分布的1.5%以内。对于含有59、79和99原子%过量15N的肽，该方法的相对标准偏差分别为4.9%、2.0和0.8%。该方法是自动化的，可以在单个LC-FTICR-MS实验中对数千个肽进行同位素富集计算。