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Aspartate Isomerization Assessment

Creative Biolabs has established a platform to predict and assess antibody degradation during the manufacturing process. Therapeutic antibodies may degrade via asparagine (Asn) deamidation and aspartate (Asp) isomerization. Aspartate isomerization is the non-enzymatic interconversion between Aspartate and Isoaspartate. In acidic environments, the peptide bond C-terminal to Aspartate is susceptible to fragmentation. Aspartate isomerization processes through intermediates similar as the Asparagine deamidation, hence the impact factors of the isomerization rate are pH, primary sequence and conformational effects.

Aspartate Isomerization

The antibody function is affected when aspartate isomerization occurs in binding area such as CDRs. Isomerization could also lead to charge heterogeneity on the protein surface and therefore results in fragmentation due to cleavage on the peptide backbone. The fragmentation reaction mostly occurs below pH 5 and Asp-Pro peptide bonds are more susceptible than other peptide bonds. Immunogenicity is potentially increased by aspartate isomerization and fragmentation leads to protein aggregation.

As the Aspartate isomerization occurs during the manufacturing process, depending on the environment conditions, the process parameters should be carefully selected to minimize the risks. For antibody candidate selection, we recommend to avoid those with potential isomerization motifs in the antigen binding interfaces where possible. The Aspartate residues with a succeeding Glycine and Proline motifs are detected for high isomerization potential. When the Aspartate isomerization cannot be tolerated at a specific position due to bioactivity or stability issues, antibody re-engineering will be performed to remove or decrease the isomerization risk. If crystal structure data is available, which allows the most complete assessment of local conformational flexibility and exposure of amino acid residues, combined with primary sequence information, Creative Biolabs guarantee to provide the most accurate prediction of aspartate isomerization.

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