Identification and molecular mechanism of action of antibacterial peptides from Flavourzyme-hydrolyzed yak casein against Staphylococcus aureus.

Antibacterial peptides can be released from yak milk casein. To date, the amino acid sequences and mechanism of action of yak casein-derived antibacterial peptides remain unknown. The current study identified antibacterial peptides from yak casein and their molecular mechanism of action. Our results showed that yak α-casein, ß-casein, and κ-casein could be effectively hydrolyzed by Flavourzyme (Solarbio Science and Technology Co. Ltd.), and the 2-h hydrolysate showed the highest antibacterial rate of 43.07 ± 2.59% against Staphylococcus aureus. The 1,000 to 3,000 Da fraction accounted for 23.61% of the 2-h hydrolysate and had an antibacterial rate of 62.64 ± 4.40%. Three novel peptides with antibacterial activity were identified from this fraction, and the ß-casein-derived peptide APKHKEMPFPKYP showed the strongest antibacterial effect (half-maximal inhibitory concentration = 0.397 mg/mL). Molecular docking predicted that APKHKEMPFPKYP interacted with 2 important enzymes of Staph. aureus, dihydrofolate reductase and DNA gyrase, through hydrophobic, hydrogen bonding, salt bridge, and π-π stacking interactions. Our findings suggest that the yak casein-derived peptides may serve as a potential source of natural preservatives to inhibit Staph. aureus.

Influence of succinylation on physicochemical property of yak casein micelles.

Succinylation is a chemical-modification method that affects the physicochemical characteristics and functional properties of proteins. This study assessed the influence of succinylation on the physicochemical properties of yak casein micelles. The results revealed that surface hydrophobicity indices decreased with succinylation. Additionally, denaturation temperature and denaturation enthalpy decreased with increasing succinylation level, except at 82%. The buffering properties of yak casein micelles were affected by succinylation. It was found that chemical modification contributed to a slight shift of the buffering peak towards a lower pH value and a markedly increase of the maximum buffering values of yak casein micelles at pH 4.5-6.0 and pH < 3. Succinylation increased yak casein micellar hydration and whiteness values. The findings obtained from this study will provide the basic information on the physicochemical properties of native and succinylated yak casein micelles.

Influence of succinylation on the conformation of yak casein micelles.

Succinylation modifies the physicochemical characteristics and improves the functional properties of proteins. This study assessed the effects of succinylation on the conformation of yak casein micelles with seven degree of modification. The results revealed that succinylation contributed to the dissociation of casein micelles. With the increase of succinylated degree, soluble nitrogen and minerals content increased, while casein micelle size and polydispersity index of micelles decreased. Succinylation affected the spatial conformation of yak casein micelles: turn decreased, ß-sheet and α-helix increased, and irregular structure were non-significantly affected. The intrinsic and ANS fluorescence intensity decreased and the maximum emission wavelength shifted red with increasing succinylation. Based on the results, the structure of yak casein micelles was characteristic of the sub-micelle model.