Antipeptide antibodies recognizing plasmin sensitive sites in bovine beta-casein sequence

Area of Science:

• Food Science
• Biochemistry
• Immunology

Background:

• Proteolysis significantly impacts cheese characteristics.
• Understanding enzymatic activity, specifically plasmin and chymosin, is crucial for cheese production.
• Bovine beta-casein is a key substrate affected by these enzymes.

Purpose of the Study:

• To develop specific antibodies for monitoring bovine beta-casein breakdown.
• To create tools for assessing plasmin and chymosin activity in cheese.
• To enable in situ monitoring of proteolysis without secondary degradation bias.

Main Methods:

• Immunization of rabbits with synthetic peptides representing plasmin-sensitive sites of beta-casein.
• Production and characterization of antisera using ACP-ELISA, Western-blot, and biosensor assays.
• In vitro hydrolysis of casein by plasmin and chymosin to validate antibody specificity.

Main Results:

• Antisera recognized beta-casein and its fragments.
• Enzymatic hydrolysis reduced determinant detection, confirming antibody specificity to cleavage sites.
• Specific antibodies detected plasmin cleavage yielding gamma1-CN and chymosin cleavage at C-terminus.

Conclusions:

• Developed antibodies are effective markers for bovine beta-casein integrity.
• Immunoassays can specifically monitor key proteolysis events in cheese.
• These tools offer a method to assess enzymatic activity without bias from secondary degradation.