A procedure for the partial fractionation of the α_s-casein complex

Abstract

A method for the partial fractionation of the a s -casein complex is described. It involves removal of K-casein from iso-electric casein by the method of Zittle & Ouster (1963) followed by fractionating the residue (a s -and /?-casein) by the urea procedure of Hipp et al. (1952). a s0 -and a sl -caseins precipitate from 3-3 M urea at pH 4-6, but much of the a s2 -, a^-and a s4 -casein remains in solution and co-precipitates with /tf-casein from 1-7 M urea at pH 4-9. Chromatography on DEAE cellulose clearly separates ^-casein from the a s2 -, a s3 -and a g4 -caseins, which are not resolved by the conditions employed. The method is suitable for the fractionation of isoelectric casein into all its major components (i) /c-casein; (ii) a s0 -and a sl -caseins; (iii) a s2 -, a s3 -and a s4 -caseins; (iv) /?-casein; (v) y-casein.Wake & Baldwin (1961) demonstrated the heterogeneity of the a s -casein fraction by electrophoresis on starch-urea gels. The proteins comprising this group had relative electrophoretic mobilities in starch or acrylamide gels of 1*16, 1-04, 1-00 and 0-86. In accordance with the recommendations of the Milk Protein Nomenclature Committee (Thompson et al. 1965) these proteins are now called a gl -, a^-, a g4 -and a g5 -casein respectively. Two additional members of the group, with relative electrophoretic mobilities of 1-19 (a s0 -) and 1-11 (a g2 -) have since been identified (ElNegoumy, 1967;Annan & Manson, 1969). The major constituent of the group, a sl -casein, was purified by and others, and has been characterized in considerable detail (Thompson et al. 1965;Rose et al. 1970). Very little work has been done on the minor a s -casein components and only recently have methods for their partial isolation been described. Annan & Manson (1969) fractionated as-casein on columns of Sulphoethyl Sephadex C-50 at pH 4-0 in the presence of 8 M urea into 3 fractions containing (i) a s0 -casein, (ii) a sl -casein and (iii) a s2 -, a s3 -and a g4 -caseins. They showed that a s3 -and a s4 -caseins were derived from a s5 -casein on reduction. Starting with the precipitate from the 50% ethanolammonium acetate purification of a sl -casein ), Hoagland et al. (1971 isolated a^-, a 84 -and a s5 -caseins by DEAE chromatography (0-01 M imidazole-HCl, 3-3 M urea, pH 7-0 buffer) and preparative gel electrophoresis. These latter authors confirmed the findings of Annan & Manson (1969) as to the origin of a g3 -and a g4 -caseins, and carried out some additional characterization studies.The present communication describes an alternative procedure for the fractionation of <x s -casein into 2 groups: (i) a s0 -and a sl -casein, (ii) a g2 -, a s3 -and a g4 -caseins. 4-2

Electrophoretic Study of Casein Breakdown during Ripening of Goat's Milk Cheese

“…Whole casein was separated by acid precipitation from skimmed goat's milk. The a.1-and 8-caseins were prepared from whole casein according to precipitation methods (Brignon et al, 1976;Fox and Guiney, 1972). Percent of total nitrogen.…”

Simultaneous Presence of PrtH and PrtH2 Proteinases in Lactobacillus helveticus Strains Improves Breakdown of the Pure α s1 -Casein

“…First, caseins were precipitated in the presence of 6.6 M urea and sulfuric acid to pH 1.5, according to the method of Zittle and Custer (47), and ␣ s -and ␤-caseins were then separated from -casein. ␣ s1 -casein was further separated from ␤-, ␥-, and ␣ s2 -caseins by dilution with urea to 3.3 M at pH 4.6, followed by ethanol precipitation according to the method of Brignon et al (2) and Fox and Guiney (12). The enriched ␣ s1 -and ␤-casein fractions were further purified by anionexchange chromatography using 210 ml of the Q Sepharose fast-flow separating phase (Pharmacia, St. Quentin Yvelines, France), according to the method of Guillou et al (19).…”

Mammals, Milk, Molecules, and Micelles

“…Three postgraduate students registered in 1970 and commenced work on (a) the isolation and characterization of pepsins (O'Leary & Fox 1973, (b) proteolysis in cheese , Phelan et al 1973), (c) fractionation of caseins (Fox & Guiney 1972), and (d ) heat stability of milk (Frank O'Mahoney, supervised by Pat Morrissey). In subsequent years, Pat took over the meat area, which he developed to a high level, and I took over work on the heat stability of milk.…”

“…I have published approximately 60 research papers (e.g., see Fox & Morrissey 1972;Fox & Guiney 1972;O'Connor & Fox 1973;McGann & Fox 1974;Mulvihill & Fox 1979a;Murphy & Fox 1991;Lucey et al 1996;O'Connell et al , 2006Malin et al 2001;Zobrist et al 2005;Huppertz et al 2006b) and 25 reviews (e.g., see , Huppertz et al 2006a, Fox & Brodkorb 2008, Uniacke-Lowe et al 2010) on milk proteins. Volume 1 of Advanced Dairy Chemistry is devoted to milk proteins.…”