Membrane glycoproteomics of fetal lung fibroblasts using LC/MS

Some aberrant N-glycosylations are being used as tumor markers, and glycoproteomicsis expected to provide novel diagnosis markers and targets of drug developments. However, one has trouble in mass spectrometric glycoproteomicsof membrane fraction because of lower intensity of glycopeptidesin the existence of surfactants. Previously, we developed a glycopeptideenrichment method by acetone precipitation, and it was successfully applied to human serum glycoproteomics. In this study, we confirmed that this method is useful to remove the surfactants and applicable to membrane glycoproteomics. The glycoproteomicapproach to the human fetal lung fibroblasts membrane fraction resulted in the identification of over 272 glycoforms on 63 sites of the 44 glycoproteins. According to the existing databases, the structural features on 41 sites are previously unreported. The most frequently occurring forms at N-glycosylation site were high- mannosetype containing nine mannoseresidues (M9) and monosialo- fucosylatedbiantennary oligosaccharides. Several unexpected N-glycans, such as fucosylatedcomplex-type and fucosylatedhigh- mannoseand/or fucosylatedpauci- mannosetypes were found in ER and lysosome proteins. Our method provides new insights into transport, biosynthesis, and degradation of glycoproteins.