Chemical and immunochemical characterization of caseins and the major whey proteins of rabbit milk

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Abstract
Caseins were separated from whey proteins by acid precipitation of skimmed rabbit milk. Whole casein was resolved by sodium dodecyl sulfate polyacrylamide-gel electrophoresis into 3 major bands with apparent relative molecular masses (Mr of 31,000, 29,000 and 25,000. On agarose/urea-gel electrophoresis whole casein gave 3 bands with electrophoretic obilities .alpha., .beta. and .gamma.. The 3 components were purified by DEAE-cellulose chromatography under denaturing and reducing conditions. Each had a different amino acid, hexose and P content, as well as non-identical peptide fragments after proteinase digestion. The 31,000 Da (dalton) protein, of .alpha.-electrophoretic mobility, had a high P content (4.38%, wt/wt); the 29,000 Da peptide, of .gamma.-mobility, had the highest hexose content (2.2%, wt/wt), contained 0.8 cysteine residue per 100 amino acid residues and was susceptible to chymosin digestion corresponding thus to .kappa.-casein; the 25,000 Da protein migrated to the .beta.-position. The rabbit casein complex is composed of at least 3 caseins, 2 of which (.alpha.- and .kappa.-caseins) are analogous to the caseins from ruminants. Although caseins are poor immunogens, specific antibodies were raised against total and purified polypeptides. The antiserum directed against whole casein recognized each polypeptide, each casein corresponding to a distinct precipitation line. The antisera directed against each casein polypeptide reacted exclusively with the coresponding casein and no antiserum cross-reaction occurred between the 3 polypeptides. From whey, several proteins were isolated, characterized and used as antigens to raise specific antibodies. An Fe-binding protein with an apparent Mr of 80,000 was immunologically and structurally identical with serum transferrin.