The Experts below are selected from a list of 225 Experts worldwide ranked by ideXlab platform
Roger J W Truscott - One of the best experts on this subject based on the ideXlab platform.
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non oxidative Modification of lens crystallins by kynurenine a novel post translational Protein Modification with possible relevance to ageing and cataract
Biochimica et Biophysica Acta, 2000Co-Authors: Brett Garner, Denis C Shaw, Robyn A Lindner, John A Carver, Roger J W TruscottAbstract:Abstract In humans, the crystallin Proteins of the ocular lens become yellow-coloured and fluorescent with ageing. With the development of senile nuclear cataract, the crystallins become brown and additional fluorophores are formed. The mechanism underlying crystallin colouration is not known but may involve interaction with kynurenine-derived UV filter compounds. We have recently identified a sulphur-linked glutathionyl-3-hydroxykynurenine glucoside adduct in the lens and speculated that kynurenine may also form adducts with GSH and possibly with nucleophilic amino acids of the crystallins (e.g. Cys). Here we show that kynurenine modifies calf lens crystallins non-oxidatively to yield coloured (365 nm absorbing), fluorescent (Ex 380 nm/Em 450–490 nm) Protein adducts. Carboxymethylation and succinylation of crystallins inhibited kynurenine-mediated Modification by approx. 90%, suggesting that Cys, Lys and possibly His residues may be involved. This was confirmed by showing that kynurenine formed adducts with GSH as well as with poly-His and poly-Lys. NMR studies revealed that the novel poly-Lys-kynurenine covalent linkage was via the ϵ-amino group of the Lys side chain and the βC of the kynurenine side chain. Analysis of tryptic peptides of kynurenine-modified crystallins revealed that all of the coloured peptides contained either His, Cys or an internal Lys residue. We propose a novel mechanism of kynurenine-mediated crystallin Modification which does not require UV light or oxidative conditions as catalysts. Rather, we suggest that the side chain of kynurenine-derived lens UV filters becomes deaminated to yield an α,β-unsaturated carbonyl which is highly susceptible to attack by nucleophilic amino acid residues of the crystallins. The inability of the lens fibre cells to metabolise their constituent Proteins results in the accumulation of coloured/fluorescent crystallins with age.
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non oxidative Modification of lens crystallins by kynurenine a novel post translational Protein Modification with possible relevance to ageing and cataract
Biochimica et Biophysica Acta, 2000Co-Authors: Brett Garner, Denis C Shaw, Robyn A Lindner, John A Carver, Roger J W TruscottAbstract:In humans, the crystallin Proteins of the ocular lens become yellow-coloured and fluorescent with ageing. With the development of senile nuclear cataract, the crystallins become brown and additional fluorophores are formed. The mechanism underlying crystallin colouration is not known but may involve interaction with kynurenine-derived UV filter compounds. We have recently identified a sulphur-linked glutathionyl-3-hydroxykynurenine glucoside adduct in the lens and speculated that kynurenine may also form adducts with GSH and possibly with nucleophilic amino acids of the crystallins (e.g. Cys). Here we show that kynurenine modifies calf lens crystallins non-oxidatively to yield coloured (365 nm absorbing), fluorescent (Ex 380 nm/Em 450-490 nm) Protein adducts. Carboxymethylation and succinylation of crystallins inhibited kynurenine-mediated Modification by approx. 90%, suggesting that Cys, Lys and possibly His residues may be involved. This was confirmed by showing that kynurenine formed adducts with GSH as well as with poly-His and poly-Lys. NMR studies revealed that the novel poly-Lys-kynurenine covalent linkage was via the epsilon-amino group of the Lys side chain and the betaC of the kynurenine side chain. Analysis of tryptic peptides of kynurenine-modified crystallins revealed that all of the coloured peptides contained either His, Cys or an internal Lys residue. We propose a novel mechanism of kynurenine-mediated crystallin Modification which does not require UV light or oxidative conditions as catalysts. Rather, we suggest that the side chain of kynurenine-derived lens UV filters becomes deaminated to yield an alpha,beta-unsaturated carbonyl which is highly susceptible to attack by nucleophilic amino acid residues of the crystallins. The inability of the lens fibre cells to metabolise their constituent Proteins results in the accumulation of coloured/fluorescent crystallins with age.
Manfred Wuhrer - One of the best experts on this subject based on the ideXlab platform.
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Linkage-specific sialic acid derivatization for MALDI-TOF-MS profiling of IgG glycopeptides.
Analytical Chemistry, 2015Co-Authors: Noortje De Haan, Karli R. Reiding, Dietmar Reusch, David Falck, Markus Haberger, Manfred WuhrerAbstract:Glycosylation is a common co- and Post-Translational Protein Modification, having a large influence on Protein properties like conformation and solubility. Furthermore, glycosylation is an important determinant of efficacy and clearance of biopharmaceuticals such as immunoglobulin G (IgG). Matrix-assisted laser desorption/ionization (MALDI)-time-of-flight (TOF)-mass spectrometry (MS) shows potential for the site-specific glycosylation analysis of IgG at the glycopeptide level. With this approach, however, important information about glycopeptide sialylation is not duly covered because of in-source and metastable decay of the sialylated species. Here, we present a highly repeatable sialic acid derivatization method to allow subclass-specific MALDI-TOF-MS analysis of tryptic IgG glycopeptides. The method, employing dimethylamidation with the carboxylic acid activator 1-ethyl-3-(3-dimethylamino)propyl)carbodiimide (EDC) and the catalyst 1-hydroxybenzotriazole (HOBt), results in different masses for the funct...
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Linkage-Specific Sialic Acid Derivatization for MALDI-TOF-MS Profiling of IgG Glycopeptides
2015Co-Authors: Noortje De Haan, Karli R. Reiding, Dietmar Reusch, David Falck, Markus Haberger, Manfred WuhrerAbstract:Glycosylation is a common co- and Post-Translational Protein Modification, having a large influence on Protein properties like conformation and solubility. Furthermore, glycosylation is an important determinant of efficacy and clearance of biopharmaceuticals such as immunoglobulin G (IgG). Matrix-assisted laser desorption/ionization (MALDI)-time-of-flight (TOF)-mass spectrometry (MS) shows potential for the site-specific glycosylation analysis of IgG at the glycopeptide level. With this approach, however, important information about glycopeptide sialylation is not duly covered because of in-source and metastable decay of the sialylated species. Here, we present a highly repeatable sialic acid derivatization method to allow subclass-specific MALDI-TOF-MS analysis of tryptic IgG glycopeptides. The method, employing dimethylamidation with the carboxylic acid activator 1-ethyl-3-(3-dimethylamino)propyl)carbodiimide (EDC) and the catalyst 1-hydroxybenzotriazole (HOBt), results in different masses for the functionally divergent α2,3- and α2,6-linked sialic acids. Respective lactonization and dimethylamidation leads to their direct discrimination in MS and importantly, both glycan and peptide moieties reacted in a controlled manner. In addition, stabilization allowed the acquisition of fragmentation spectra informative with respect to glycosylation and peptide sequence. This was in contrast to fragmentation spectra of underivatized samples, which were dominated by sialic acid loss. The method allowed the facile discrimination and relative quantitation of IgG Fc sialylation in therapeutic IgG samples. The method has considerable potential for future site- and sialic acid linkage-specific glycosylation profiling of therapeutic antibodies, as well as for subclass-specific biomarker discovery in clinical IgG samples derived from plasma
Patricio Yankilevich - One of the best experts on this subject based on the ideXlab platform.
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global site specific neddylation profiling reveals that neddylated cofilin regulates actin dynamics
Nature Structural & Molecular Biology, 2020Co-Authors: Annette M Vogl, Lilian Phu, Raquel Becerra, Sebastian A Giusti, Erik Verschueren, Trent Hinkle, Martin Diego Bordenave, Max Adrian, Amy Heidersbach, Patricio YankilevichAbstract:Neddylation is the Post-Translational Protein Modification most closely related to ubiquitination. Whereas the ubiquitin-like Protein NEDD8 is well studied for its role in activating cullin−RING E3 ubiquitin ligases, little is known about other substrates. We developed serial NEDD8-ubiquitin substrate profiling (sNUSP), a method that employs NEDD8 R74K knock-in HEK293 cells, allowing discrimination of endogenous NEDD8- and ubiquitin-Modification sites by MS after Lys-C digestion and K-eGG-peptide enrichment. Using sNUSP, we identified 607 neddylation sites dynamically regulated by the neddylation inhibitor MLN4924 and the de-neddylating enzyme NEDP1, implying that many non-cullin Proteins are neddylated. Among the candidates, we characterized lysine 112 of the actin regulator cofilin as a novel neddylation event. Global inhibition of neddylation in developing neurons leads to cytoskeletal defects, altered actin dynamics and neurite growth impairments, whereas site-specific neddylation of cofilin at K112 regulates neurite outgrowth, suggesting that cofilin neddylation contributes to the regulation of neuronal actin organization. NEDD8-ubiquitin substrate profiling (sNUSP) identifies neddylation sites in many non-cullin Proteins. Among the candidates, neddylation of cofilin regulates actin dynamics and neurite growth and outgrowth in developing neurons.
David Wendehenne - One of the best experts on this subject based on the ideXlab platform.
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S-nitrosylation: an emerging Post-Translational Protein Modification in plants.
Plant Science, 2011Co-Authors: Jeremy Astier, Christian Lindermayr, Sumaira Rasul, Emmanuel Koen, Hamid Manzoor, Angélique Besson-bard, Olivier Lamotte, Sylvain Jeandroz, Joerg Durner, David WendehenneAbstract:Increasing evidences support the assumption that nitric oxide (NO) acts as a physiological mediator in plants. Understanding its pleiotropic effects requires a deep analysis of the molecular mechanisms underlying its mode of action. In the recent years, efforts have been made in the identification of plant Proteins modified by NO at the Post-Translational level, notably by S-nitrosylation. This reversible process involves the formation of a covalent bond between NO and reactive cysteine residues. This research has now born fruits and numerous Proteins regulated by S-nitrosylation have been identified and characterized. This review describes the basic principle of S-nitrosylation as well as the Biotin Switch Technique and its recent adaptations allowing the identification of S-nitrosylated Proteins in physiological contexts. The impact of S-nitrosylation on the structure/function of selected Proteins is further discussed.
Brett Garner - One of the best experts on this subject based on the ideXlab platform.
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non oxidative Modification of lens crystallins by kynurenine a novel post translational Protein Modification with possible relevance to ageing and cataract
Biochimica et Biophysica Acta, 2000Co-Authors: Brett Garner, Denis C Shaw, Robyn A Lindner, John A Carver, Roger J W TruscottAbstract:Abstract In humans, the crystallin Proteins of the ocular lens become yellow-coloured and fluorescent with ageing. With the development of senile nuclear cataract, the crystallins become brown and additional fluorophores are formed. The mechanism underlying crystallin colouration is not known but may involve interaction with kynurenine-derived UV filter compounds. We have recently identified a sulphur-linked glutathionyl-3-hydroxykynurenine glucoside adduct in the lens and speculated that kynurenine may also form adducts with GSH and possibly with nucleophilic amino acids of the crystallins (e.g. Cys). Here we show that kynurenine modifies calf lens crystallins non-oxidatively to yield coloured (365 nm absorbing), fluorescent (Ex 380 nm/Em 450–490 nm) Protein adducts. Carboxymethylation and succinylation of crystallins inhibited kynurenine-mediated Modification by approx. 90%, suggesting that Cys, Lys and possibly His residues may be involved. This was confirmed by showing that kynurenine formed adducts with GSH as well as with poly-His and poly-Lys. NMR studies revealed that the novel poly-Lys-kynurenine covalent linkage was via the ϵ-amino group of the Lys side chain and the βC of the kynurenine side chain. Analysis of tryptic peptides of kynurenine-modified crystallins revealed that all of the coloured peptides contained either His, Cys or an internal Lys residue. We propose a novel mechanism of kynurenine-mediated crystallin Modification which does not require UV light or oxidative conditions as catalysts. Rather, we suggest that the side chain of kynurenine-derived lens UV filters becomes deaminated to yield an α,β-unsaturated carbonyl which is highly susceptible to attack by nucleophilic amino acid residues of the crystallins. The inability of the lens fibre cells to metabolise their constituent Proteins results in the accumulation of coloured/fluorescent crystallins with age.
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non oxidative Modification of lens crystallins by kynurenine a novel post translational Protein Modification with possible relevance to ageing and cataract
Biochimica et Biophysica Acta, 2000Co-Authors: Brett Garner, Denis C Shaw, Robyn A Lindner, John A Carver, Roger J W TruscottAbstract:In humans, the crystallin Proteins of the ocular lens become yellow-coloured and fluorescent with ageing. With the development of senile nuclear cataract, the crystallins become brown and additional fluorophores are formed. The mechanism underlying crystallin colouration is not known but may involve interaction with kynurenine-derived UV filter compounds. We have recently identified a sulphur-linked glutathionyl-3-hydroxykynurenine glucoside adduct in the lens and speculated that kynurenine may also form adducts with GSH and possibly with nucleophilic amino acids of the crystallins (e.g. Cys). Here we show that kynurenine modifies calf lens crystallins non-oxidatively to yield coloured (365 nm absorbing), fluorescent (Ex 380 nm/Em 450-490 nm) Protein adducts. Carboxymethylation and succinylation of crystallins inhibited kynurenine-mediated Modification by approx. 90%, suggesting that Cys, Lys and possibly His residues may be involved. This was confirmed by showing that kynurenine formed adducts with GSH as well as with poly-His and poly-Lys. NMR studies revealed that the novel poly-Lys-kynurenine covalent linkage was via the epsilon-amino group of the Lys side chain and the betaC of the kynurenine side chain. Analysis of tryptic peptides of kynurenine-modified crystallins revealed that all of the coloured peptides contained either His, Cys or an internal Lys residue. We propose a novel mechanism of kynurenine-mediated crystallin Modification which does not require UV light or oxidative conditions as catalysts. Rather, we suggest that the side chain of kynurenine-derived lens UV filters becomes deaminated to yield an alpha,beta-unsaturated carbonyl which is highly susceptible to attack by nucleophilic amino acid residues of the crystallins. The inability of the lens fibre cells to metabolise their constituent Proteins results in the accumulation of coloured/fluorescent crystallins with age.