A simplified method for the purification of riboflavin-binding protein from hen's egg yolk

Murthy, Usha S.; Sreekrishna, K.; Adiga, P. Radhakantha

doi:10.1016/0003-2697(79)90668-7

Cited by 19 publications

(6 citation statements)

References 19 publications

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“…Despite the fact that anisotropic broadening of the signals decreases the spectral resolution.v-" valuable information about proton hyperfine couplings can be obtained. 3 is obtained after removal of riboflavin and reconstitution with lumiflavin deuterated at the 8-methyl group (5). As can be deduced from Table 2, the HFC constants of methyl protons in 7, 8 and 10 positions (including the anisotropic components of the methyl groups in 8 and 10 positions) and that of the a-proton in position 6 could be determined.…”

Section: Resultsmentioning

confidence: 99%

¹H and ²H ENDOR Investigations of Flavin Radicals Bound to Riboflavin Binding Protein from Egg White

Bretz

Henzel

Kurreck

et al. 1989

Israel Journal of Chemistry

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Riboflavin‐binding apoprotein isolated from egg white has been used as a protein matrix for a variety of substituted and/or deuterated flavins. The neutral radical states of the reconstituted flavoproteins were studied by ENDOR (electron nuclear double resonance) spectroscopy in disordered solids. For the first time all major hyperfine coupling constants of a flavin cofactor embedded within a protein could be measured and unambiguously assigned to molecular positions. Some conclusions about the geometrical arrangement of the flavin within the protein pocket and about molecular motions could be drawn.

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Section: Resultsmentioning

confidence: 99%

¹H and ²H ENDOR Investigations of Flavin Radicals Bound to Riboflavin Binding Protein from Egg White

Bretz

Henzel

Kurreck

et al. 1989

Israel Journal of Chemistry

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“…Apo-RBPY was isolated and purified according to Murthy et al (1979) with the exception that an Amicon concentrator was used instead of aquacide treatment. Apo-RBPW was isolated and purified according to Farrell et al (1969).…”

Section: Methodsmentioning

confidence: 99%

Carbon-13 and nitrogen-15 nuclear magnetic resonance study on the interacton between riboflavin and riboflavin-binding apoprotein

Moonen¹,

Berg²,

Boerjan³

et al. 1984

Biochemistry

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“…The conventional procedures of lipid removal, such as extraction with diethyl ether (Morton, 1955) or butan-1-ol (Meslar & White, 1979), are cumbersome and are often unsatisfactory, either owing to incomplete extraction at low temperatures, resulting in subsequent colloid formation, or to significant protein denaturation at elevated temperatures. To circumvent these difficulties, an alternate, efficient, yet mild, procedure adopted in the present investigation stems from a previous observation (Raju & Mahadevan, 1975;Murthy et al, 1979) that the lipoproteins are almost entirely excluded from DEAE-cellulose and the water-soluble proteins firmly held on the ion-exchanger are easily recovered by elution with 1 M-NaCl. This exclusionchromatographic step afforded nearly 10-fold enrichment of the water-soluble yolk proteins.…”

Section: Discussionmentioning

confidence: 99%

“…The sources of other chemicals and biochemicals have been detailed previously (Muniyappa & Adiga, 1979). Apo-(riboflavin-binding protein) from the egg yolk was purified by the procedure developed previously (Murthy et al, 1979).…”

Section: Methodsmentioning

confidence: 99%

Nature of the thiamin-binding protein from chicken egg yolk

Muniyappa¹,

Adiga²

1981

Biochemical Journal

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A simple, rapid and efficient procedure for the purification of thiamin-binding protein from chicken egg yolk was developed. The method involved removal, by exclusion, of lipoproteins from DEAE-cellulose and subsequent elution of water-soluble proteins held on the ion-exchanger with 1 M-NaCl, followed by treatment of the eluted protein fraction with an aqueous suspension of dextran/charcoal to generate apoprotein from the holoprotein. The resultant protein fraction was subjected to bioaffinity chromatography on thiamin pyrophosphate--AE (aminoethyl)-Sepharose. The protein eluted specifically with 10 microM-thiamin at pH 7.0, was homogeneous by the criteria of polyacrylamide-gel disc electrophoresis, had a mol.wt. of 38 000 +/- 2000 and was not a glycoprotein. The purified thiamin-binding protein specifically interacted with riboflavin-binding protein with no detectable deleterious affect on its (14C)thiamin-binding capacity. The protein bound [14C]thiamin with a molar ratio of 1.0, with dissociation constant (Kd) 0.41 microM. This protein-ligand interaction was inhibited by thiamin analogues and antagonists. The absorption spectrum of the protein in the presence of thiamin exhibited significant hypochromism at the 278 nm band, indicating the involvement of aromatic amino acid residues of the protein, during its binding to the ligand. The protein cross-reacted with the monospecific antiserum to egg-white thiamin-binding protein, showing thereby that thiamin-binding proteins present in chicken egg yolk and white are the products of the same structural gene.

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A simplified method for the purification of riboflavin-binding protein from hen's egg yolk

Cited by 19 publications

References 19 publications

¹H and ²H ENDOR Investigations of Flavin Radicals Bound to Riboflavin Binding Protein from Egg White

¹H and ²H ENDOR Investigations of Flavin Radicals Bound to Riboflavin Binding Protein from Egg White

Carbon-13 and nitrogen-15 nuclear magnetic resonance study on the interacton between riboflavin and riboflavin-binding apoprotein

Nature of the thiamin-binding protein from chicken egg yolk

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A simplified method for the purification of riboflavin-binding protein from hen's egg yolk

Cited by 19 publications

References 19 publications

1H and 2H ENDOR Investigations of Flavin Radicals Bound to Riboflavin Binding Protein from Egg White

1H and 2H ENDOR Investigations of Flavin Radicals Bound to Riboflavin Binding Protein from Egg White

Carbon-13 and nitrogen-15 nuclear magnetic resonance study on the interacton between riboflavin and riboflavin-binding apoprotein

Nature of the thiamin-binding protein from chicken egg yolk

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Product

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¹H and ²H ENDOR Investigations of Flavin Radicals Bound to Riboflavin Binding Protein from Egg White

¹H and ²H ENDOR Investigations of Flavin Radicals Bound to Riboflavin Binding Protein from Egg White