biostudies-literatureLoke ICancer Institute NSW1507-1527https://www.ebi.ac.uk/biostudies/studies/S-EPMC5546201biostudies-literatureUnknown16(8)Human neutrophil elastase (HNE) is an important <i>N</i>-glycosylated serine protease in the innate immune system, but the structure and immune-modulating functions of HNE <i>N</i>-glycosylation remain undescribed. Herein, LC-MS/MS-based glycan, glycopeptide and glycoprotein profiling were utilized to first determine the heterogeneous <i>N</i>-glycosylation of HNE purified from neutrophil lysates and then from isolated neutrophil granules of healthy individuals. The spatiotemporal expression of HNE during neutrophil activation and the biological importance of its <i>N</i>-glycosylation were also investigated using immunoblotting, cell surface capture, native MS, receptor interaction, protease inhibition, and bacteria growth assays. Site-specific HNE glycoprofiling demonstrated that unusualMolecular & cellular proteomics : MCPPaucimannose-Rich <i>N</i>-glycosylation of Spatiotemporally Regulated Human Neutrophil Elastase Modulates Its Immune Functions.PMC554620113/ECF/1–02Packer NHOstergaard OHeegaard NHHLoke IThaysen-Andersen MfalsePaucimannose-Rich <i>N</i>-glycosylation of Spatiotemporally Regulated Human Neutrophil Elastase Modulates Its Immune Functions.Human neutrophil elastase (HNE) is an important <i>N</i>-glycosylated serine protease in the innate immune system, but the structure and immune-modulating functions of HNE <i>N</i>-glycosylation remain undescribed. Herein, LC-MS/MS-based glycan, glycopeptide and glycoprotein profiling were utilized to first determine the heterogeneous <i>N</i>-glycosylation of HNE purified from neutrophil lysates and then from isolated neutrophil granules of healthy individuals. The spatiotemporal expression of HNE during neutrophil activation and the biological importance of its <i>N</i>-glycosylation were also investigated using immunoblotting, cell surface capture, native MS, receptor interaction, protease inhibition, and bacteria growth assays. Site-specific HNE glycoprofiling demonstrated that unusual2017-01-01T00:00:00Z2017 Aug2026-05-03T03:48:51.984Z2019-03-26T23:48:39ZS-EPMC55462012863008710.1074/mcp.M116.06674610.1074/mcp.m116.066746