Probable Identification of γ, TS-, R- and S-Caseins as Fragments of β-Casein

Gordon, William G.; Groves, M.L.; Greenberg, Rae; Jones, Susan B.; Kalan, Edwin B.; Peterson, Robert F.; Townend, Robert

doi:10.3168/jds.s0022-0302(72)85469-9

Cited by 61 publications

(20 citation statements)

References 7 publications

Supporting

Mentioning

Contrasting

Unclassified

Order By: Relevance

“…The method also gives a reasonably satisfactory separation of the minor y o y 2 -and y 3 -caseins from other caseins, but does not separate them completely from each other. The latter is not a serious drawback since these materials are closely related, probably being derived from /?-casein (Gordon et al 1972;Groves et al 1973), and for most compositional studies it will suffice to treat them as a single fraction. K-Casein, on the other hand, is only partly resolved by the present system since only about 80% of it is recovered as pure material (fraction 2a), with the remainder being present in fraction 2b together with about an equal amount of some other proteins.…”

Section: Quantitative Analysis Of Fractions Obtained By Chromatographymentioning

confidence: 98%

“…The patterns for fractions la-4b are shown in Plate 1 (slots la-4b respectively) together with that for the alkylated whole casein (slot AC), while the corresponding patterns for the samples after chymosin treatment are shown in Plate 2, with parts A and B of the latter plate showing the materials which migrated at pH 7-9 towards the anode and cathode respectively. The patterns enabled all fractions, apart from fraction 2b, to be identified readily and indicated that fraction la consisted almost entirely of y 2 -casein (TS-casein), fraction lb of y 3 -casein (R-casein) together possibly with y r casein (y-casein), fraction 2a of K-casein, fraction 3 of /?-casein, fraction 4a -in order of increasing mobility -of the minor a s -caseins a g6 , a s4 , a s3 and Os 2 , and fraction 4b of a sl -and a s0 -caseins (Thompson, 1966;Groves & Gordon, 1969;Groves et al 1973;Annan & Manson, 1969). The nomenclature adopted for the y-caseins is that recommended by Whitney et al (1976).…”

Section: Qualitative Analysis Of Fractions Obtained By Chromatographymentioning

confidence: 99%

See 1 more Smart Citation

An improved method for the quantitative fractionation of casein mixtures using ion-exchange chromatography

Davies

Law

1977

Journal of Dairy Research

107

View full text Add to dashboard Cite

A method for the quantitative analysis of casein mixtures is described. The method uses ion-exchange chromatography at 4 °C on DEAE cellulose, Whatman DE52 in the presence of tris-chloride-urea buffer (pH 8-6), and a NaCl gradient to fractionate the alkylated casein mixtures and a micro-biuret technique to determine protein concentrations. Values for fractions representative of (1) y-caseins, (2) /c-casein plus some unknown proteins, (3) /?-casein, (4) minor a g -caseins and (5) a 8l -plus a 80 -caseins were obtained, and whole casein from herd bulk milk was found to consist on average of about 2, 13, 36, 11 and 38% respectively of these fractions. Agreement between duplicate fractionations was satisfactory and the recovery of material varied between 94 and 98%.Rose, Davies & Yaguchi (1969) described a method for the quantitative determination of the major components of casein mixtures which was based on the use of column chromatography on DEAE cellulose. The method gave a more extensive fractionation than that obtained by moving-boundary electrophoresis (Rolleri, Larson & Touchberry, 1956;Larson & Kendall, 1957), and yielded results which were more accurate than those derived from sialic acid (Marier, Tessier & Rose, 1963) and turbidimetric measurements (Tessier, Rose & Marier 1963) and also probably more reliable than those obtained from scanning patterns obtained by gel electrophoresis (Morr, Lin & Josephson, 1971). In the method of Rose et al. fractionation was done on DEAE cellulose, Whatman DE11, an anion exchanger with low proteinbinding capacity and relatively poor kinetic properties which has now been superseded by more advanced ion-exchange materials. In an attempt to improve on their method the improved exchanger Whatman DE52 has been used for casein fractionation and certain aspects of the procedure, such as the methods used for protein determination and for the measurement of the recovery of material, have been more critically examined. These studies have produced a system giving greater resolution of casein mixtures than achieved previously and have shown that agreement between duplicate fractionations and recovery of material is satisfactory. However, complete separation of /c-casein from other proteins was not possible and values for /c-casein are therefore somewhat overestimated.

show abstract

Section: Quantitative Analysis Of Fractions Obtained By Chromatographymentioning

confidence: 98%

Section: Qualitative Analysis Of Fractions Obtained By Chromatographymentioning

confidence: 99%

An improved method for the quantitative fractionation of casein mixtures using ion-exchange chromatography

Davies

Law

1977

Journal of Dairy Research

107

View full text Add to dashboard Cite

show abstract

“…The specificity of plasmin on p -casein is well documented (Gordon et al, 1972;Eigel, 1977Eigel, , 1981 and its specificity on a S 2-casein has been determined recently (Le Bars and Gripon, 1989a, b;Visser et al, 1989). However, it appears that the specificity of plasmin on bovine cm-casein has not been reported and was the subject of this study.…”

Section: Introductionmentioning

confidence: 95%

Proteolytic specificity of plasmin on bovine α_s1‐Casein

et al. 1993

View full text Add to dashboard Cite

Proteolytic specificity of plasmin on bovine α s1 -Casein, Food Biotechnology, 7:2, 143-158, ABSTRACTThe proteolytic specificity of plasmin (fibrinolysin, E. C. 3.4.21.7, from bovine plasma) on bovine α s1 -casein was determined in solution in 50 mM ammonium bicarbonate buffer, pH 8.4, at 37°C.Peptides, isolated by reverse-phase high performance liquid chromatography on a C 18 column or by electroblotting from urea-polyacrylamide gel electrophoretograms, were identified from their amino acid sequence and mass. The principal plasmin cleavage sites were found at Arg 22 -Phe 23 , Arg 90 -Tyr 91 , Lys 102 -Lys 103 , Lys 103 -Tyr 104 , Lys 105 -Val 106 , Lys 124 -Glu 125 and Arg 151 -Gln 152 .The initial cleavage sites and the order of production of small (pH 4.6-soluble) peptides suggest that α s1 -casein was cleaved initially towards the centre of the molecule.*Present address:

show abstract

“…Results of amino acid analyses, molecular weight determinations, peptide mappings, and end-group analyses first indicated a close relationship between several minor casein fractions (yI-, 72-, and y3-casein) and f3-casein (9). Comparison of the amino acid sequence of the first 10 to 15 residues at the NH2 terminus of each of these minor caseins with the primary structure reported for /-casein revealed that 'y-, Y2-, and y3-caseins are actually identical to residues 28-209, 106-209, and of f-casein, respectively (8,9). Eigel (10) demonstrated formation of yr-, 72-, and -y3-caseins by incubation of bovine ,B-casein with plasmin.…”

mentioning

confidence: 99%

Plasmin-mediated proteolysis of casein in bovine milk.

Eigel¹,

Hofmann²,

Chibber³

et al. 1979

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

109

View full text Add to dashboard Cite

Plasminogen was found to be present in bovine milk by crossreactivity between rabbit antiserum to plasminogen and casein prepared from milk by acid precipitation. This result was further supported by recovery of intact 125I-labeled plasminogen from rabbit milk after its intravenous injection. Freshly isolated whole bovine casein was observed to undergo slow autoproteolysis at 37°C. Polyacrylamide gel electrophoresis revealed gradual disappearance of major caseins accompanied by appearance and increase in intensity of numerous electrophoretic bands. This autoproteolysis was inhibited by low concentrations of &aminocaproic acid (0.1 mM) and diisopropyl fluorophosphate (1 mM); catalytic amounts of urokinase accelerated the process. Autoproteolysis of isolated bovine ,B-casein was shown by both urea and sodium dodecyl sulfate gel electrophoresis to result, in formation of Irl-and -Y2-caseins. Similar electrophoretic bands were formed when V-asein was degraded by plasmin prepared from bovine blood serum. These results support the hypothesis that bovine plasmin occurs in milk and is identical to alkaline milk protease.Plasminogen is the zymogen of the proteolytic enzyme plasmin which is responsible for the dissolution of fibrin clots in blood. In humans and other species several organs have been shown to contain considerable amounts of plasminogen activator activity (1-3). However, plasminogen itself is normally considered to be a constituent only of mammalian blood plasma (4). The presence of a naturally occurring protease in milk was first reported in 1897 (5). Casein, prepared by acid precipitation of skim milk, contains most of the proteolytic activity found in milk (6). Kaminogawa et al. (7) Immunological Study. Antiserum to bovine plasminogen was raised by weekly subcutaneous injection of 1 ml of plasminogen (10 mg) in phosphate-buffered saline containing 50% Freund's complete adjuvant into a New Zealand White rabbit. Five weeks after the first injection, blood was collected by cardiac puncture, allowed to coagulate for 30 min at 23°C, and centrifuged at 1500 X g for 10 min (16). The globulin fraction was prepared by two successive precipitations at 50% ammonium sulfate saturation (17) followed by washing with phosphate-buffered saline. After extensive dialysis against distilled water at 4°C, the globulin fraction was recovered by lyophili- Rabbit plasminogen was prepared as described above. Sodium dodecyl sulfate (NaDodSO4) gel electrophoresis of rabbit plasminogen was conducted in 7.5% acrylamide slabs (20) and gels were stained with 0.05% Coomassie blue. Gels were destained electrophoretically, dehydrated under reduced pressure, and exposed to x-ray film for 7 days.Abbreviations: EACA, E-aminocaproic acid; iPr2P-F, diisopropyl fluorophosphate; UKase, urokinase; NaDodSO4, sodium dodecyl sulfate. 2244 The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S. C. §1734 solely to indicat...

show abstract

Probable Identification of γ, TS-, R- and S-Caseins as Fragments of β-Casein

Cited by 61 publications

References 7 publications

An improved method for the quantitative fractionation of casein mixtures using ion-exchange chromatography

An improved method for the quantitative fractionation of casein mixtures using ion-exchange chromatography

Proteolytic specificity of plasmin on bovine α_s1‐Casein

Plasmin-mediated proteolysis of casein in bovine milk.

Contact Info

Product

Resources

About

Probable Identification of γ, TS-, R- and S-Caseins as Fragments of β-Casein

Cited by 61 publications

References 7 publications

An improved method for the quantitative fractionation of casein mixtures using ion-exchange chromatography

An improved method for the quantitative fractionation of casein mixtures using ion-exchange chromatography

Proteolytic specificity of plasmin on bovine αs1‐Casein

Plasmin-mediated proteolysis of casein in bovine milk.

Contact Info

Product

Resources

About

Proteolytic specificity of plasmin on bovine α_s1‐Casein