Micelle Stability: κ-Casein Structure and Function

Abstract

The stability of the casein micelle is dependent on the presence of κ-casein (CN) on the surface of the micelle where it functions as an interface between the hydrophobic caseins of the micelle interior and the aqueous environment. κ-Casein is also involved in thiol-catalyzed disulfide interchange reactions with the whey proteins during heat treatments and, after rennet cleavage, in the facilitation of micelle coagulation. These functions of κ-CN are regulated by the three-dimensional structure of the protein on the micelle surface. The usual means of determining structure are not available for κ-CN because this protein is strongly self-associating and has never been crystallized. Instead, algorithms were used to predict selected secondary structures and circular dichroism spectroscopy on κ-CN and the macropeptide released by chymosin. Three peptides were synthesized to cover the chymosin-sensitive site (His98-Lys111), the region in the macropeptide that could be helical (Pro130-Ile153), and the region between. Nuclear magnetic resonance spectroscopy showed that the peptide His98-Lys111 was probably a β-strand with tight turns at each end. This hypothesis was confirmed by a study of the molecular dynamics showing that the C variant of κ-CN interacted less strongly with chymosin; consequently, the slow renneting time of milk that contains this protein was explainable. Both circular dichroism and nuclear magnetic resonance indicated that the peptide Pro130-Ile153 was probably helical under normal physiological conditions. A preliminary study using nuclear magnetic resonance showed that the intervening peptide had no discernible secondary structure. Consequently, most of the β-sheet structure of κ-CN is likely in the para-κ-CN region.