Year: 1968
Type: article
Abstract: A procedure for the preparation of pure K-casein by Sephadex gel filtration is described.Unreduced casein is filtered through a 2.5-by 95-em colmnn of Sephadex G-150 equilibrated with 0.005 ~ triscitrate buffer, pit 8.6, containing 6 urea.K-Casein is excluded from the gel and emerges as a sharp peak at the void volume; all other caseins are retarded by the gel.The method is particularly suitable for preparing pure Kicasein in good yield from small quantities of whole casein, but larger amounts of n-casein are readily prepared.The method is also suitable for the purification of K-casein-rich preparations.In milk mieelles, and in preparations such as acid-precipitated whole casein, K-casein forms a complex with a~-a~d fl-caseins.Urea dissociates this complex (8), the a~-and fl-caseins being converted into monomeric forms while Kcasein remains as relatively large aggregates with intermoleeular disulfide bonding (4, 6, 13).Yaguehi and Tarassuk (18) observed that when a solution of acid casein at pH 8.6 passed through a column of Sephadex G-100, which had been equilibrated with 6 ~ urea at pH 8.6, relatively pure K-casein was eluted at the void volume of column.Also, Nakai et al. (5) report that K-casein was eluted solely at the void volume when unreduced n-casein was filtered through columns of Sephadex G-200 equilibrated with either tris-hydrochlorie acidurea (pH 7.6) or acetic acid-urea, or ureasodium chloride, solutions.These obselwations suggested that gel filtration nfight prove useful in the preparation of n-casein.The present paper presents a method for the routine preparation of pure n-casein from acid casein, or from n-casein-rich materials, which is convenient and produces an undegraded product with little or no contaminating protein. Methods and MaterialsPreparation of whole caseins.Various fresh skinmfilks from individual cows and fresh herd milk were used, but all data reported in detail were obtained in tests with milk, from a single Received for publication September 26, 1967.N.R.C. no.9981.Ayrshire cow, containing as-casein B, fl-casein A, and K-casein A.Acid casein was prepared at pH 4.6 and washed with water.Soluble casein was prepared according to yon Ihippel and Waugh (15), except that the micellar casein was brought into solution by adding a mixture of di-and tetrasodium salts of ethylenediaminetetraacetic acid (EDTA) to chelate the calcium.
Source: Journal of Dairy Science
Authors: Makoto Yaguchi, D.T. Davies, Y.K. Kim
Institution: National Research Council Canada
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: bronze
OpenAlex ID: W2018787174