Journal of Dairy Research

Table of Contents - February 1996 - Volume 63, Issue 01  

Original Articles

Comparative study of methods for the isolation and purification of bovine κ-casein and its hydrolysis by chymosin

Kate P. Coolbeara1†, David F. Elgara1, Tim Coolbeara2 and John S. Ayersa1

a1 Department of Chemistry and Biochemistry, Massey University, Private Bag 11222, Palmerston North, New Zealand

a2 New Zealand Dairy Research Institute, Private Bag 11029, Palmerston North, New Zealand

Summary

κ-Casein was purified from a single batch of whole acid casein (κ-A variant) using different methods in order to compare their merits in producing a purified material with a carbohydrate and phosphate heterogeneity representative of the whole κ-casein complement in milk. Ion-exchange methods of purification gave products of higher purity than precipitation techniques involving final purification by ethanol fractionation, but all methods resulted in κ-caseins of apparently similar heterogeneity and chemical composition. The purified κ-caseins were hydrolysed with chymosin and the derived macropeptides isolated. These were all virtually identical as determined by reversed-phase chromatography and gel electrophoresis. Some observations on chymosin hydrolysis of κ-casein were made. In addition to formation of the major para-κ-casein (Glu1–Phe105) and macropeptide (Met106–Val169), chymosin hydrolysis at pH 6·6 also resulted in two minor para-κ-caseins with N-termini corresponding to Phe18 and Ser33 of κ-casein. At pH 5·5 and 4·5 para-κ-casein was rapidly hydrolysed into at least six fragments, one of which had an N-terminus corresponding to Trp76 of κ-casein. At pH 6·6, 5·5 and 4·5 the κ-casein macropeptide was stable to chymosin, but at pH 2·3 it was hydrolysed by chymosin into fragments with N-termini corresponding to Met106, He125, Ala138, Val139, Thr145 and Glu147 of κ-casein.

(Received November 29 1994)

(Accepted June 08 1995)

Footnotes

† For correspondence.