Modifications in Protein Structure and Xanthine Oxidase Inhibition of Yak Casein Induced by Protease Treatment

ABSTRACT Yak milk is abundant in casein, which can generate a variety of bioactive peptides through enzymatic hydrolysis. However, the influence of enzymatic hydrolysis on the structural properties of yak casein and its inhibitory effects on xanthine oxidase (XOD) remain largely unexplored. This study demonstrated that when yak casein was subjected to treatment with flavourzyme, the degree of hydrolysis progressively increased over time, resulting in the fragmentation of the casein's flake‐like structure into smaller particles. Circular dichroism analysis revealed that after 4 h of enzymatic treatment, there was an elevation in the β‐sheet content of the yak casein hydrolysate, while other secondary structure elements diminished. Furthermore, flavourzyme treatment induced modifications in the tertiary structure of yak casein. The study also examined the impact of varying hydrolysis durations on XOD inhibitory activity, discovering that the hydrolysate obtained after 3 h displayed the highest inhibition on XOD, with an inhibition rate of (40.63 ± 3.36) %. Additionally, the fraction of the hydrolysate with a molecular weight exceeding 3 kDa demonstrated enhanced XOD inhibitory activity. This study is the first to investigate how varying hydrolysis durations with flavourzyme affect the structural characteristics of yak casein and its XOD inhibitory activity.