Dataset Information

Glycoproteomic analysis of Haloferax volcanii strains H53, DaglB, and Dagl15

ABSTRACT: An in-depth glycoproteomic analysis of Haloferax volcanii has been performed, comparing the wildtype H53 with knockout mutants of aglB and agl15. While AglB-dependent glycosylation has been described to occur under standard culture conditions, the Agl15-dependent glycosylation pathway has previously been described to be only active under low-salt conditions. In this dataset, the largest archaeal glycoproteome could be identified with more than 40 N-glycoproteins and more than 100 N-glycopptides. It reveals not only that both glycosylation pathways can be active under the same conditions but also that the same glycosites can be modified by different glycans from both glycosylation pathways.

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Haloferax Volcanii (halobacterium Volcanii)

SUBMITTER: Stefan Schulze

LAB HEAD: Mechthild Pohlschroder

PROVIDER: PXD021874 | Pride | 2021-06-20

REPOSITORIES: Pride

ACCESS DATA

Dataset's files

Source:

			Action	DRS
	20190321_StS_H53_Cyt_GluC_stepped.raw	Raw
	20190321_StS_H53_Cyt_Try_stepped.raw	Raw
	20190321_StS_H53_Mem_GluC_stepped.raw	Raw
	20190321_StS_H53_Mem_Try_stepped.raw	Raw
	20190321_StS_H53_SN_GluC_stepped.raw	Raw

Items per page:

1 - 5 of 67

Publications

Comprehensive glycoproteomics shines new light on the complexity and extent of glycosylation in archaea.

Schulze Stefan S Pfeiffer Friedhelm F Garcia Benjamin A BA Pohlschroder Mechthild M

PLoS biology 20210617 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 th ...[more]

PMID: 34138841

Similar Datasets

Project description:Recombinant proteins are of great interest in glycobiology and proteomics, known especially for their reproducibility and accessibility. However, variation in glycosylation among recombinant glycoproteins is not well understood and may depend on numerous conditions in the biomanufacturing process. In order to confidently assess variation in glycosylation measurements, it is vital to both optimize the measurement of, and determine the degree of variation between, distributions of glycosylation on specific sites of glycoproteins. This is especially important for glycoproteins that are known to have rapid sequence changes, such as with different influenza strains. In this study, eight strains of recombinant influenza hemagglutinin and neuraminidase produced from HEK293 cell line were obtained from four vendors and digestion was conducted using a series of complex multi-enzymatic methods designed to isolate glycopeptide sequons. Site-specific glycosylation profiles of intact glycopeptides were produced using mass spectrometric evaluation on an orbitrap system and compared using spectral similarity scores. Variation in glycan abundances and distribution was most pronounced between different strains of virus (similarity score = 383 out of 1000), whereas replicates resulted in low variation (similarity score = 957 out of 1000). Glycan variation was also measured based on differences between vendors, lots, batches, protease digestion, and intra-protein site. The most abundant glycans in all of these influenza glycoproteins were monofucosylated and complex, as reported by other laboratories. However, it was found that different vendors can produce very different glycan distributions for the same glycosylation site. Notably, it is demonstrated that glycan distributions are similar for conserved regions of influenza glycoproteins. Overall, these methods present a potential use in developing reproducible measurements of glycosylated biologics for quality control or making more informed decisions in biomanufacturing.