Toward a framework for sulfoproteomics: Synthesis and characterization of sulfotyrosine‐containing peptides

Seibert, Christoph; Sakmar, Thomas P.

doi:10.1002/bip.20821

Cited by 105 publications

(170 citation statements)

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“…Thus, we decided to investigate further the mechanism by which nitration of phenylalanine and Tyr residues can lead to a cross-reactive antibody response to native proteins. Specifically, we demonstrate that autoantibody responses are elicited when mice are immunized with TNF-α mutants containing pNO 2 Phe and 3NO 2 Tyr, as well as with sulfotyrosine (SO 3 Tyr) residues, a PTM involved in the regulation of chemokine signaling (25,26). Mechanistic studies indicate that, although CD4 T cells remain tolerant to endogenous epitopes, neoepitopes activate CD4 T cells that render help for the generation of autoantibody responses.…”

mentioning

confidence: 93%

Loss of CD4 T-cell–dependent tolerance to proteins with modified amino acids

Gauba

Grünewald

Gorney

et al. 2011

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

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The site-specific incorporation of the unnatural amino acid p-nitrophenylalanine (pNO 2 Phe) into autologous proteins overcomes self-tolerance and induces a long-lasting polyclonal IgG antibody response. To determine the molecular mechanism by which such simple modifications to amino acids are able to induce autoantibodies, we incorporated pNO 2 Phe, sulfotyrosine (SO 3 Tyr), and 3-nitrotyrosine (3NO 2 Tyr) at specific sites in murine TNF-α and EGF. A subset of TNF-α and EGF mutants with these nitrated or sulfated residues is highly immunogenic and induces antibodies against the unaltered native protein. Analysis of the immune response to the TNF-α mutants in different strains of mice that are congenic for the H-2 locus indicates that CD4 T-cell recognition is necessary for autoantibody production. IFN-γ ELISPOT analysis of CD4 T cells isolated from vaccinated mice demonstrates that peptides with mutated residues, but not the wild-type residues, are recognized. Immunization of these peptides revealed that a CD4 repertoire exists for the mutated peptides but is lacking for the wild-type peptides and that the mutated residues are processed, loaded, and presented on the I-A b molecule. Overall, our results illustrate that, although autoantibodies are generated against the endogenous protein, CD4 cells are activated through a neo-epitope recognition mechanism. Therefore, tolerance is maintained at a CD4 level but is broken at the level of antibody production. Finally, these results suggest that naturally occurring posttranslational modifications such as nitration may play a role in antibody-mediated autoimmune disorders.loss of tolerance | autologous antibody response | protein nitration P reviously we showed that the site-specific incorporation of the unnatural amino acid p-nitrophenylalanine (pNO 2 Phe) into a protein can induce autoantibodies against the endogenous, unmodified protein (1, 2). The murine proteins TNF-α and retinolbinding protein (RBP4) were modified genetically to incorporate pNO 2 Phe at several surface-exposed positions and were used as immunogens. Vaccination of mice with these altered proteins resulted in an IgG polyclonal antibody response against the endogenous proteins that lasted for at least 40 wk. Experiments involving hybridomas generated from the TNF-α-immunized mice revealed that the IgG antibody response was directed against different regions of the protein and not exclusively against the region containing the pNO 2 Phe residue. The autoantibodies generated by the mutated proteins were tested in an endotoxemia model for their ability to neutralize wild-type TNF-α. Protection from endotoxemia indicated that vaccination with pNO 2 Phemodified self-protein is able to elicit physiologically relevant autoantibody responses. Although these initial studies implicated a role for CD4 T cells in autoantibody production, the exact mechanism by which the site-specific incorporation of pNO 2 Phe breaks immunological self-tolerance remained to be elucidated.Recent studies have indicated that postt...

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mentioning

confidence: 93%

Loss of CD4 T-cell–dependent tolerance to proteins with modified amino acids

Gauba

Grünewald

Gorney

et al. 2011

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

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Section: Introductionmentioning

confidence: 99%

“…Modification is detected on Tyr (mainly), Ser and Thr residues and has the same nominal mass increase as phosphorylation (+80 Da) [8][9][10]. Thus, it was suggested that rates of protein sulfonation could be underestimated due to a coexistence of both phosphorylated and sulfonated (isobaric) forms of the same peptide [8][9][10]. Use of ultra-high resolution (accurate mass) measurements, ultraviolet and infrared photodissociation spectroscopy techniques demonstrated that PTM modified isobaric peptides could be distinguished [11][12][13].…”

Section: Introductionmentioning

confidence: 99%

Study of the gas-phase fragmentation behaviour of sulfonated peptides

Škulj

Rožman

2015

International Journal of Mass Spectrometry

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a b s t r a c tA series of singly and doubly protonated peptides bearing sulfonated residue have been studied, using both experiment and molecular modelling, to elucidate fragmentation chemistry of sulfonated peptides. Collision-induced dissociation mass spectra indicate that the sulfo group loss (neutral loss of 80 Da) is the dominant dissociation channel. Modelling results suggest the proton transfer mechanism, where upon vibrational excitation, the acidic side chain proton is transferred from the sulfo group hydroxyl to the ester oxygen resulting in S O bond cleavage and formation of the unmodified hydroxyl containing residue and SO 3 . Conformations associated with potential energy profile of the reaction imply the charge remote nature of the proposed mechanism. The proposed proton transfer mechanism was compared with the intramolecular nucleophilic substitution (S N 2) mechanism, the main pathway suggested for neutral loss of phosphoric acid from phosphopeptides. Both pathways (proton transfer and S N 2) are available for sulfonated and phosphorylated peptides; however, each posttranslational modification favours different mechanism. The change of the bond dissociation enthalpies and the ability of stabilising the transition state structures are demonstrated as main factors responsible for each posttranslational modification activating a different pathway.

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“…Protein Tyr sulfation, which is catalyzed by tyrosylprotein sulfotransferases (TPSTs), is believed to be critical for the activating of these peptides. TPSTs were first identified in mouse and human (Ouyang et al, 1998) and have been shown to play important roles in many physiological and pathological processes, including hormonal regulation, hemostasis, inflammation, and infectious diseases (Seibert and Sakmar, 2008). Interestingly, while TPST orthologs from numerous vertebrate and invertebrate species were readily identifiable based on sequence homology, no plant TPST ortholog was identified this way, suggesting that plant TPSTs have evolved in a manner distinct from their animal counterparts (Moore, 2009).…”

Section: Introductionmentioning

confidence: 99%

ArabidopsisTyrosylprotein Sulfotransferase Acts in the Auxin/PLETHORA Pathway in Regulating Postembryonic Maintenance of the Root Stem Cell Niche

et al. 2010

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Recent identification of the Arabidopsis thaliana tyrosylprotein sulfotransferase (TPST) and a group of Tyr-sulfated peptides known as root meristem growth factors (RGFs) highlights the importance of protein Tyr sulfation in plant growth and development. Here, we report the action mechanism of TPST in maintenance of the root stem cell niche, which in the Arabidopsis root meristem is an area of four mitotically inactive quiescent cells plus the surrounding mitotically active stem cells. Mutation of TPST leads to defective maintenance of the root stem cell niche, decreased meristematic activity, and stunted root growth. We show that TPST expression is positively regulated by auxin and that mutation of this gene affects auxin distribution by reducing local expression levels of several PIN genes and auxin biosynthetic genes in the stem cell niche region. We also show that mutation of TPST impairs basal-and auxin-induced expression of the PLETHORA (PLT) stem cell transcription factor genes and that overexpression of PLT2 rescues the root meristem defects of the loss-offunction mutant of TPST. Together, these results support that TPST acts to maintain root stem cell niche by regulating basal-and auxin-induced expression of PLT1 and PLT2. TPST-dependent sulfation of RGFs provides a link between auxin and PLTs in regulating root stem cell niche maintenance.

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Toward a framework for sulfoproteomics: Synthesis and characterization of sulfotyrosine‐containing peptides

Cited by 105 publications

References 154 publications

Loss of CD4 T-cell–dependent tolerance to proteins with modified amino acids

Loss of CD4 T-cell–dependent tolerance to proteins with modified amino acids

Study of the gas-phase fragmentation behaviour of sulfonated peptides

ArabidopsisTyrosylprotein Sulfotransferase Acts in the Auxin/PLETHORA Pathway in Regulating Postembryonic Maintenance of the Root Stem Cell Niche

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