Mass Spectrometric Determination of the Sites of O-Glycan Attachment with Low Picomolar Sensitivity

Rademaker, Geert Jan; Pergantis, Spiros A.; Blok-Tip, Leonore; Langridge, James; Kleen, Astrid; Thomas‐Oates, Jane

doi:10.1006/abio.1997.2548

Cited by 108 publications

(91 citation statements)

References 29 publications

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“…Finally, it is of interest to note that chemical base-catalyzed ␤-elimination has been used extensively in combination with mass spectrometry for the analysis of serine and threonine phosphorylation (29 -31) and O-glycosylation (32)(33)(34). Therefore, we attempted to employ the same strategy on our synthetic sulfopeptides to ascertain whether this approach might be useful for determination of sulfonation sites.…”

Section: Resultsmentioning

confidence: 99%

O-Sulfonation of Serine and Threonine

Medzihradszky

Darula

Perlson

et al. 2004

Molecular & Cellular Proteomics

130

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Protein sulfonation on serine and threonine residues is described for the first time. This post-translational modification is shown to occur in proteins isolated from organisms representing a broad span of eukaryote evolution, including the invertebrate mollusk Lymnaea stagnalis, the unicellular malaria parasite Plasmodium falciparum, and humans. Detection and structural characterization of this novel post-translational modification was carried out using liquid chromatography coupled to electrospray tandem mass spectrometry on proteins including a neuronal intermediate filament and a myosin light chain from the snail, a cathepsin-C-like enzyme from the parasite, and the cytoplasmic domain of the human orphan receptor tyrosine kinase Ror-2. These findings suggest that sulfonation of serine and threonine may be involved in multiple functions including protein assembly and signal transduction. Molecular & Cellular Proteomics 3:429 -443, 2004.Sulfonation occurs as a common enzymatic modification of endogenous substances including proteins, carbohydrates, catecholamines, and estrogenic steroids as well as xenobiotic chemicals (1). Sulfonation refers to the transfer of the sulfonate group (SO 3 Ϫ1 ) from 3Ј-phosphoadenosine-5Ј-phosphosulfate (PAPS), 1 the only known sulfonate donor (2), and can occur through several types of linkages, such as esters and anhydrides (O-sulfonation), amides (N-sulfonation), and thioesters (S-sulfonation), of which O-sulfonation is the most prominent (3). The transfer of SO 3 Ϫ1 to a hydroxyl or phenolic acceptor (O-sulfonation) generates a sulfono-derivative, and this reaction has commonly been referred to as sulfation rather than the more accurate O-sulfonation. The majority of cellular sulfonation is of the O type and occurs primarily on polysaccharides, steroids, catecholamines, and thyroid hormones (1). These reactions are catalyzed by the soluble cytosolic sulfotransferases and appear to alter their bioactivity. For example, estrogen, testosterone, and thyroid hormones (T 3 and T 4 ) can interact with their respective receptors to regulate transcription, whereas their sulfate-containing moieties cannot. Furthermore, the half-life of these compounds in blood is significantly shorter than that of their conjugated counterparts, suggesting that sulfonation maintains these compounds in an inactive state ready for rapid deployment by the removal of the sulfonyl group.While the cytosolic sulfotransferases conjugate cell-permeable or intracellular compounds, the membrane-bound Golgiassociated sulfotransferases are primarily responsible for sulfonation of extracellular proteins via a co-or post-translational mechanism. The membrane-bound sulfotransferases are responsible for the sulfonation of various glycosaminoglycans, such as heparin and heparan sulfate. Additionally, these enzymes catalyze the direct sulfonation of proteins on the O 4 position of tyrosine residues (4). It is one of the last modifications to occur during protein transiting the trans-Golgi and thus has been found almost ex...

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Section: Resultsmentioning

confidence: 99%

O-Sulfonation of Serine and Threonine

Medzihradszky

Darula

Perlson

et al. 2004

Molecular & Cellular Proteomics

130

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“…Proteins were solubilized in sample buffer according to the manufacturer's instructions and resolved by isoelectric focusing on the precast IEF strips followed by SDS-PAGE on homogenous 12.5% slab gels, 20 ϫ 20 cm, for the second dimension. Gels were stained with Bio-Safe colloidal G-250 Coomassie Blue stain (Bio-Rad Laboratories) or silver-stained (20). For subsequent lectin probing, the gels were electroblotted onto PVDF membrane at 50 V for 1 h in 10 mM 3-(cyclohexylamino)-1-propanesulfonic acid buffer, pH 11, containing 10% methanol.…”

Section: Methodsmentioning

confidence: 99%

Structure of the N-Linked Glycan Present on Multiple Glycoproteins in the Gram-negative Bacterium, Campylobacter jejuni

Young¹,

Brisson

Kelly

et al. 2002

Journal of Biological Chemistry

398

396

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“…Because we have shown above that it was possible to detach the sugar from the protein by ISD, we sought to observe the oxonium ion of the sugar and fragment it to get insight into its structure (26,27). This was done by using a Q-TOF Premier (Waters, Milford, MA) mass spectrometer, which allows both mass accuracy and efficient fragmentation for low mass ions.…”

Section: Collision-induced Sugar Fragmentation Using Quadrupole Tof (mentioning

confidence: 99%

Alternative Neisseria spp. type IV pilin glycosylation with a glyceramido acetamido trideoxyhexose residue

Chamot‐Rooke

Rousseau

Lanternier

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

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The importance of protein glycosylation in the interaction of pathogenic bacteria with their host is becoming increasingly clear. Neisseria meningitidis, the etiological agent of cerebrospinal meningitis, crosses cellular barriers after adhering to host cells through type IV pili. Pilin glycosylation genes (pgl) are responsible for the glycosylation of PilE, the major subunit of type IV pili, with the 2,4-diacetamido-2,4,6-trideoxyhexose residue. Nearly half of the clinical isolates, however, display an insertion in the pglBCD operon, which is anticipated to lead to a different, unidentified glycosylation. Here the structure of pilin glycosylation was determined in such a strain by ''top-down'' MS approaches. MALDI-TOF, nanoelectrospray ionization Fourier transform ion cyclotron resonance, and nanoelectrospray ionization quadrupole TOF MS analysis of purified pili preparations originating from N. meningitidis strains, either wild type or deficient for pilin glycosylation, revealed a glycan mass inconsistent with 2,4-diacetamido-2,4,6-trideoxyhexose or any sugar in the databases. This unusual modification was determined by in-source dissociation of the sugar from the protein followed by tandem MS analysis with collision-induced fragmentation to be a hexose modified with a glyceramido and an acetamido group. We further show genetically that the nature of the sugar present on the pilin is determined by the carboxyl-terminal region of the pglB gene modified by the insertion in the pglBCD locus. We thus report a previously undiscovered monosaccharide involved in posttranslational modification of type IV pilin subunits by a MS-based approach and determine the molecular basis of its biosynthesis. mass spectrometry ͉ pglB ͉ Pili

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Mass Spectrometric Determination of the Sites of O-Glycan Attachment with Low Picomolar Sensitivity

Cited by 108 publications

References 29 publications

O-Sulfonation of Serine and Threonine

O-Sulfonation of Serine and Threonine

Structure of the N-Linked Glycan Present on Multiple Glycoproteins in the Gram-negative Bacterium, Campylobacter jejuni

Alternative Neisseria spp. type IV pilin glycosylation with a glyceramido acetamido trideoxyhexose residue

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