Self-association of casein studied using enzymatic cross-linking at different temperatures. (April 2019) - Raak, Norbert
Abstract: Sodium caseinate and β-casein self-associate to casein particles in solution because of hydrophobic interactions. Microbial transglutaminase (mTGase) presumably preferentially cross-links molecules that are located within the same particles, meaning that polymer size is limited to the number of monomers initially present in the particles. The aim of this study was to affect the self-association of casein by varying the temperature and thereby controlling the maximum size of casein polymers formed using mTGase. Activity and stability of mTGase were determined at different temperatures, showing that about 10 times more enzyme had to be added at 10 than at 40 °C to compensate for differences in enzyme activity. Analysis with gel electrophoresis showed that incubation temperature had no effect on maximum polymer size at the selected protein concentration (27 g/kg), resulting in similar stiffness of acid-induced gels after incubation for 24 h. For a more detailed characterization of casein polymers, size exclusion chromatography was coupled to multi-angle light scattering (MALS). Estimated molar mass distributions of casein polymers were similar at both incubation temperatures and the increase in molar mass leveled off after moderate incubation time. This underlines the idea of a maximum polymer size and suggests no cross-linking between existing polymers. Besides that, the MALS detector showed a contaminant of low concentration but large size which co-eluted with casein polymers and possibly led to overestimation of their molar masses. Highlights: Sodium caseinate and β-casein were cross-linked using microbial transglutaminase (mTGase). Activity and stability of mTGase were determined as a function of temperature. Incubation temperature had no effect on maximum polymer size. Size exclusion chromatography was coupled to multi-angle light scattering. Molar mass determination confirmed the occurrence of maximum polymer sizes.