LTQ数据关联中性丢失扫描鉴定糖基化肽段

简介:

Glycosylation is an important post-translational modification associated with many proteins that have a regulatory function. Several LC-MS/MS approaches have been used for the analysis and structural elucidation of glycoproteins. Most commonly, a glycoprotein is enzymatically digested, resulting in fragments that are fractionated by reversed phase-LC. The peptide fractions may be analyzed by either on-line MS/MS, or collected and analyzed offline by MALDI. Peptides that do not correspond to predicted masses may represent glycosylated forms. These peptides are subsequently treated with a glycosidase to cleave off the oligosaccharide. The difference in the mass following a cleavage is used to infer the carbohydrate constituents. Generally, this method does not allow determination of the oligosaccharide structure or of the exact site of its attachment to the peptide. Glycoproteins can be challenging to analyze because they are generally present in low concentration in cells. In addition, glycopeptides are often hydrophilic and do not bind well to reversedphase columns commonly used in peptide analysis, making determination of the position of elution difficult. This report describes a method for using a Data Dependent Neutral Loss experiment and the high sensitivity MSn capabilities of the Thermo Scientific LTQ linear ion trap mass spectrometer to characterize glycoproteins.

仪器:

LTQ ion trap mass spectrometer

结论:

These results demonstrate that the ultra-high sensitivity and high spectral quality offered by the LTQ mass spectrometer make it ideal for the analysis of enzymatically modified glycoproteins. The Data Dependent Neutral Loss experiment presented here enables determination of the position of elution, whereas the high sensitivity MS/MS and MS3 spectra generate information-rich data for structural elucidation of the various glycoforms of ribonuclease B.