biostudies-literatureSchulze SDeutsche ForschungsgemeinschaftNIAID NIH HHSFoundation for the National Institutes of HealthNCI NIH HHSNational Science Foundatione3001277https://www.ebi.ac.uk/biostudies/studies/S-EPMC8241124biostudies-literatureUnknown19(6)Glycosylation is one of the most complex posttranslational protein modifications. Its importance has been established not only for eukaryotes but also for a variety of prokaryotic cellular processes, such as biofilm formation, motility, and mating. However, comprehensive glycoproteomic analyses are largely missing in prokaryotes. Here, we extend the phenotypic characterization of N-glycosylation pathway mutants in Haloferax volcanii and provide a detailed glycoproteome for this model archaeon through the mass spectrometric analysis of intact glycopeptides. Using in-depth glycoproteomic datasets generated for the wild-type (WT) and mutant strains as well as a reanalysis of datasets within the Archaeal Proteome Project (ArcPP), we identify the largest archaeal glycoproteome described so far.PLoS biologyComprehensive glycoproteomics shines new light on the complexity and extent of glycosylation in archaea.PMC82411243986254471817518P01 CA196539AI118891R01 AI118891Schulze SPohlschroder MPfeiffer FGarcia BAfalseComprehensive glycoproteomics shines new light on the complexity and extent of glycosylation in archaea.Glycosylation is one of the most complex posttranslational protein modifications. Its importance has been established not only for eukaryotes but also for a variety of prokaryotic cellular processes, such as biofilm formation, motility, and mating. However, comprehensive glycoproteomic analyses are largely missing in prokaryotes. Here, we extend the phenotypic characterization of N-glycosylation pathway mutants in Haloferax volcanii and provide a detailed glycoproteome for this model archaeon through the mass spectrometric analysis of intact glycopeptides. Using in-depth glycoproteomic datasets generated for the wild-type (WT) and mutant strains as well as a reanalysis of datasets within the Archaeal Proteome Project (ArcPP), we identify the largest archaeal glycoproteome described so far.2021-01-01T00:00:00Z2021 Jun2026-05-07T22:28:59.161Z2025-06-01T00:34:15.509ZS-EPMC82411243413884110.1371/journal.pbio.3001277