Deglycosylation with trifluoromethanesulfonic acid differentially affects inhibitor activities of turkey ovomucoid

Dabich, Danica; Yurewicz, Edward C.; Battel, Virginia A.

doi:10.1016/0167-4838(93)90110-d

Cited by 3 publications

(5 citation statements)

References 32 publications

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“…The 98.4% carbohydrate removal from OM as measured spectrophotometrically was even higher than that described by Gu et al (1989) and similar to that reported by Dabich et al (1993), who removed about 75 and 94% by TFMS treatment respectively. Since hydrolysis with TFMS does not release GlcNAc residues attached to Asn (Edge et al, 1981;Tarns & Welinder, 1995) the five isoforms of d-OM measured by MALDI-TOF may be due to uncleaved GlcNAc.…”

Section: Discussionsupporting

confidence: 84%

“…For example, Gu et al (1989) found no change in trypsin inhibitory and immunological activity of hen's egg OM after chemical deglycosylation, but suggested that the carbohydrate moieties contribute to the stability of OM against tryptic hydrolysis and heat denaturation. In contrast, Dabich et al (1993) observed decreasing trypsin inhibitory activity of deglycosylated OM from turkey's egg. The specific interaction of carbohydrate epitopes in IgE binding has been described for different allergens.…”

Section: Introductionmentioning

confidence: 47%

“…Both results are in agreement with data reported for SDS-PAGE and physicochemical methods respectively (Burley & Vedehra, 1989;Ebbeh0j et al, 1995). Overestimation of the MW for OM and d-OM from turkeys' eggs was observed in SDS-PAGE by Dabich et al (1993). The lower electrophoretic mobility of OM may be due to a lower SDS/protein ratio since carbohydrate moieties do not bind SDS.…”

Section: Discussionmentioning

confidence: 87%

“…It was confirmed that OA and OM are frequent allergens in egg-white. The applied TFMS deglycosylation is reported to hydrolyze glycosidic linkages without affecting peptide bonds (Edge et al, 1981;Dabich et al, 1993). Complete deglycosylation of OM was assumed by a decrease in MW as determined by SDS-PAGE and MALDI-TOF mass spectrometry.…”

Section: Discussionmentioning

confidence: 99%

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Allergenicity of hen's egg‐white proteins: IgE binding of native and deglycosylated ovomucoid

Besler¹,

Steinhart²,

Paschke³

1997

Food and Agricultural Immunology

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Ovomucoid (OM) is a major allergen of hen's egg-white. Carbohydrate moieties from the glycoprotein were removed by chemical deglycosylation. Deglycosylated ovomucoid (d-OM) consists of five isoforms with molecular weights (MW) between 20.7 and 21.5 kDa whereas native OM has a MW of 28 kDa as determined by matrix-assisted laser desorption/ionization time of flight mass spectrometry. The IgE-binding properties of both proteins were investigated. All patients' sera tested, showed strong IgE binding to both native OM and d-OM in SDS-PAGE/immunoblot. In enzyme-allergosorbent test experiments, 50% inhibition of IgE binding was observed, at almost the same inhibitor concentrations of 67 and 57 ng ml -1 for native OM and d-OM respectively. Results indicate that only epitopes on the protein backbone are responsible for IgE binding while carbohydrate residues do not participate in allergenic structures of OM.

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Section: Discussionsupporting

confidence: 84%

Section: Introductionmentioning

confidence: 47%

Section: Discussionmentioning

confidence: 87%

Section: Discussionmentioning

confidence: 99%

See 2 more Smart Citations

Allergenicity of hen's egg‐white proteins: IgE binding of native and deglycosylated ovomucoid

Besler¹,

Steinhart²,

Paschke³

1997

Food and Agricultural Immunology

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“…The glycoprotein without carbohydrate was degraded by excess trypsin [116] unlike the native molecule, indicating that glycosylation of this inhibitor is imperative for protecting the protein inhibitor from its target protease. Dabich et al [117] found that turkey ovomucoid was rendered less effective as a trypsin inhibitor, an activity dependent on the second domain of the molecule, but had increased activity towards chymotrypsin, an activity that resides in the third domain, but may be constrained by glycosylation of the second domain. The cysteine protease inhibitor thiostatin depended on glycosylation for its inhibitory activity [118], but retained its reactivity with antibodies to the native protein.…”

Section: Structure-function Relationships In Glycoproteinsmentioning

confidence: 99%

Deglycosylation of glycoproteins with trifluoromethanesulphonic acid: elucidation of molecular structure and function

EDGE

2003

Biochemical Journal

114

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The alteration of proteins by post-translational modifications, including phosphorylation, sulphation, processing by proteolysis, lipid attachment and glycosylation, gives rise to a broad range of molecules that can have an identical underlying protein core. An understanding of glycosylation of proteins is important in clarifying the nature of the numerous variants observed and in determining the biological roles of these modifications. Deglycosylation with TFMS (trifluoromethanesulphonic acid) [Edge, Faltynek, Hof, Reichert, and Weber, (1981) Anal. Biochem. 118, 131-137] has been used extensively to remove carbohydrate from glycoproteins, while leaving the protein backbone intact. Glycosylated proteins from animals, plants, fungi and bacteria have been deglycosylated with TFMS, and the most extensively studied types of carbohydrate chains in mammals, the N-linked, O-linked and glycosaminoglycan chains, are all removed by this procedure. The method is based on the finding that linkages between sugars are sensitive to cleavage by TFMS, whereas the peptide bond is stable and is not broken, even with prolonged deglycosylation. The relative susceptibility of individual sugars in glycosidic linkage varies with the substituents at C-2 and the occurrence of amido and acetyl groups, but even the most stable sugars are removed under conditions that are sufficiently mild to prevent scission of peptide bonds. The post-translational modifications of proteins have been shown to be required for diverse biological functions, and selective procedures to remove these modifications play an important role in the elucidation of protein structure and function.

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Structurally unique recombinant Kazal-type proteinase inhibitor retains activity when terminally extended and glycosylated

Kłudkiewicz

Kodrı́k

Grzelak

et al. 2005

Protein Expression and Purification

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Deglycosylation with trifluoromethanesulfonic acid differentially affects inhibitor activities of turkey ovomucoid

Cited by 3 publications

References 32 publications

Allergenicity of hen's egg‐white proteins: IgE binding of native and deglycosylated ovomucoid

Allergenicity of hen's egg‐white proteins: IgE binding of native and deglycosylated ovomucoid

Deglycosylation of glycoproteins with trifluoromethanesulphonic acid: elucidation of molecular structure and function

Structurally unique recombinant Kazal-type proteinase inhibitor retains activity when terminally extended and glycosylated

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