Hoffman Michael D, Sniatynski Matthew J, Rogalski Jason C, Le Blanc J C Yves, Kast Juergen
The Biomedical Research Centre, University of British Columbia, Vancouver, British Columbia, Canada.
J Am Soc Mass Spectrom. 2006 Mar;17(3):307-17. doi: 10.1016/j.jasms.2005.11.002. Epub 2006 Jan 27.
Post-translational modifications of proteins are involved in determining the activity of proteins and are essential for proper protein function. Current mass spectrometric strategies require one to specify a particular type of modification, in some cases also a particular charge state of a protein or peptide that is to be studied before the actual analysis. Due to these requirements, most of the modifications on proteins are not considered in such an experiment and, thus, a series of similar analyses need to be performed to ensure a more extensive characterization. A novel scan strategy has been developed, multiple neutral loss monitoring (MNM), allowing for the comprehensive screening of post-translational modifications (PTM) on proteins that fragment as neutral losses in a mass spectrometer. MNM method parameters were determined by performing product ion scans on a number of modified peptides over a range of collision energies, providing neutral loss energy profiles and optimal collision energies (OCE) for each modification, supplying valuable information pertaining to the fragmentation of these modifications and the necessary parameters that would be required to obtain the best analysis. As the optimal collision energy was highly dependent on the type of modification and the charge state of the peptide, the MNM scan was operated with a collision energy gradient. Autocorrelation analyses identified the type of modification, and convolution mapping analyses identified the associated peptide. The MNM scan with the new collision energy parameters was successfully applied to a mixture of four modified peptides in a BSA digest. The implementation of this technique will allow for comprehensive screening of all modifications that fragment as neutral losses.
蛋白质的翻译后修饰参与决定蛋白质的活性，对于蛋白质的正常功能至关重要。当前的质谱分析策略要求在实际分析之前指定特定类型的修饰，在某些情况下还需指定要研究的蛋白质或肽段的特定电荷状态。由于这些要求，此类实验中大多不考虑蛋白质上的大多数修饰，因此需要进行一系列类似分析以确保更广泛的表征。已开发出一种新型扫描策略，即多中性丢失监测（MNM），可对在质谱仪中以中性丢失形式断裂的蛋白质上的翻译后修饰（PTM）进行全面筛选。通过在一系列碰撞能量下对多个修饰肽段进行产物离子扫描来确定MNM方法参数，从而为每种修饰提供中性丢失能量分布和最佳碰撞能量（OCE），提供与这些修饰的断裂有关的有价值信息以及获得最佳分析所需的必要参数。由于最佳碰撞能量高度依赖于修饰类型和肽段的电荷状态，MNM扫描以碰撞能量梯度运行。自相关分析确定修饰类型，卷积映射分析确定相关肽段。具有新碰撞能量参数的MNM扫描已成功应用于牛血清白蛋白消化物中四种修饰肽段的混合物。该技术的实施将允许对以中性丢失形式断裂的所有修饰进行全面筛选。
