Complete structure elucidation of a globular protein by particle beam liquid chromatography-Fourier transform infrared spectrometry and electrospray liquid chromatography-mass spectrometry sequence and conformation of β-lactoglobulin

Turula, Vincent E.; Bishop, Randall T.; Ricker, Robert D.; Haseth, James A. de

doi:10.1016/s0021-9673(96)00960-0

Cited by 17 publications

(6 citation statements)

References 10 publications

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“…The observed decrease in the fluorescence intensity related with heat denaturation as shown in this study is likely to be related to self-quenching of the fluorescence occurring when two identical fluorescent molecules are in close proximity to each other (25). On the other hand, proteolytic degradation such as trypsination splits the aggregates of β-lg and hydrolyzes it, resulting in 19 peptide fragments containing 1-26 amino acids (30). Increasing the distance between the adjacent fluorescent dye molecules, by proteolysis, decreases their interaction and thus increases their fluorescence intensity (25).…”

Section: Discussionmentioning

confidence: 52%

Enterocyte and M-Cell Transport of Native and Heat-Denatured Bovine β-Lactoglobulin: Significance of Heat Denaturation

Rytkönen

Valkonen²,

Virtanen³

et al. 2006

J. Agric. Food Chem.

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The three-dimensional structure, digestibility, and immunological properties of bovine beta-lactoglobulin (beta-lg) are modified by heat treatments used in processing of liquid milk products. Because it is not known if such treatments also modify the intestinal transport properties of beta-lg, the transport of native and heat-denatured bovine beta-lg was investigated in experimental cell models using Caco-2 cells and M cells. Transport of beta-lg labeled with a fluorescent marker was followed with fluorometric measurements, electrophoretic analyses, and fluorescence microscopy. The data show that both cell types transported native beta-lg more efficiently than they did heat-denatured beta-lg. In addition, M cells transported native beta-lg more than Caco-2 cells. Transport of native and heat-denatured beta-lg was transcellular. The electrophoretic data also suggest that heat-denatured beta-lg may have degraded more than native beta-lg during the transport.

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Section: Discussionmentioning

confidence: 52%

Enterocyte and M-Cell Transport of Native and Heat-Denatured Bovine β-Lactoglobulin: Significance of Heat Denaturation

Rytkönen

Valkonen²,

Virtanen³

et al. 2006

J. Agric. Food Chem.

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“…Variant B is effectively the D64G/V118A double mutant of variant A (31). There are 15 Lys, 2 His and 1 free Cys residues in both variants of β-LG.…”

Section: Resultsmentioning

confidence: 99%

“…β-LG is composed of two variants, a heavier A chain and a lighter B chain. Variant B is effectively the D64G/V118A double mutant of variant A (31). There are 15 Lys, two His, and one free Cys residues in both variants of β-LG.…”

Section: Resultsmentioning

confidence: 99%

Covalent Cross-Linking of Glutathione and Carnosine to Proteins by 4-Oxo-2-nonenal

Zhu

Gallogly

Mieyal

et al. 2009

Chem. Res. Toxicol.

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The lipid oxidation product 4-oxo-2-nonenal (ONE) derived from peroxidation of polyunsaturated fatty acids is a highly reactive protein cross-linking reagent. The major family of cross-links reflects conjugate addition of side-chain nucleophiles such as sulfhydryl or imidazole groups to the C=C of ONE to give either a 2- or 3-substituted 4-ketoaldehyde, which then undergoes Paal-Knorr condensation with the primary amine of protein lysine side-chains. If ONE is intercepted in biological fluids by antielectrophiles such as glutathione (GSH) or β-alanylhistidine (carnosine), this would lead to circulating 4-ketoaldehydes that could then bind covalently to the protein Lys residues. This phenomenon was investigated by SDS–PAGE and mass spectrometry (MALDI-TOF and LC-ESI-MS/MS with both tryptic and chymotryptic digestion). Under the reaction conditions of 0.25 mM to 2 mM ONE, 1 mM GSH or carnosine, 0.25 mM bovine β-lactoglobulin (β-LG), 100 mM phosphate buffer (pH 7.4, 10% ethanol), 24 h, 37 °C, virtually every Lys of β-LG was found to be fractionally cross-linked to GSH. Cross-linking of Lys to carnosine was slightly less efficient. Using cytochrome c and RNase A, we showed that ONE becomes more protein-reactive in the presence of GSH, whereas protein modification by 4-hydroxy-2-nonenal is inhibited by GSH. Stable antielectrophile–ONE–protein cross-links may serve as biomarkers of oxidative stress and may represent a novel mechanism of irreversible protein glutathionylation.

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“…ESI‐MS played important role in the complete structure elucidation of BLG [22]. Covalent adducts of BLG with acrylamide [23] and non‐covalent complexes with 8‐anilinonaphthalene‐1‐sulfonic acid [23], 4‐hydroxy‐2‐nonenal [24] and different pesticides [25] were also reported.…”

Section: Introductionmentioning

confidence: 99%

Induced chirality upon binding of cis‐parinaric acid to bovine β‐lactoglobulin: spectroscopic characterization of the complex

Zsila¹,

Imre²,

Szabó³

et al. 2002

FEBS Letters

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Binding of the polyunsaturated cis-parinaric acid to bovine L L-lactoglobulin (BLG) was studied by circular dichroism (CD), electronic absorption spectroscopy and mass spectrometry methods. Upon protein binding, the UV absorption band of parinaric acid is red shifted by ca. 5 nm, showing hypochromism and reduced vibrational fine structure, suggesting that the ligand binds as a monomer in non-planar geometry. In the CD spectra measured at pH 7.36 and 8.5 a strong, negative Cotton band appears centered at 310 nm (v vO O = 3 325 M 31 cm 31 ) corresponding to the long-wavelength absorption band of cis-parinaric acid. The source of this induced optical activity is the helical distortion of the polyene chromophore caused by the chiral protein environment. From CD spectral data the value of the association constant was calculated to be 4.7U U10 5 M 31 at pH 7.36. CD and mass spectrometry measurements showed that parinaric acid binds weakly to BLG in acidic solution, though small peaks at mass 18559 and 18645 can be obtained in the reconstructed electrospray mass spectrum; these correspond to the binding of parinaric acid in 1:1 stoichiometry to both monomer variants of BLG B and A. The hydrophobic interior cavity of BLG was assigned as the primary binding site of cisparinaric acid. ß 2002 Federation of European Biochemical Societies. Published by Elsevier Science B.V. All rights reserved.

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Complete structure elucidation of a globular protein by particle beam liquid chromatography-Fourier transform infrared spectrometry and electrospray liquid chromatography-mass spectrometry sequence and conformation of β-lactoglobulin

Cited by 17 publications

References 10 publications

Enterocyte and M-Cell Transport of Native and Heat-Denatured Bovine β-Lactoglobulin: Significance of Heat Denaturation

Enterocyte and M-Cell Transport of Native and Heat-Denatured Bovine β-Lactoglobulin: Significance of Heat Denaturation

Covalent Cross-Linking of Glutathione and Carnosine to Proteins by 4-Oxo-2-nonenal

Induced chirality upon binding of cis‐parinaric acid to bovine β‐lactoglobulin: spectroscopic characterization of the complex

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