Computational engineering of Protein L ligands to achieve an optimal a nity resin for purification of antibody fragments

Protein L a nity chromatography can be a well-established platform for the purification of antibody fragments containing kappa light chain in biopharmaceutical downstream processing. A nity chromatography resins usually suffer from low binding capacity and this problem increases the cost of the final product at the industrial production. Recent studies have shown that an increase in the binding a nity of resin ligands towards their target proteins leads to increased product purity, recovery and dynamic binding capacity values in a nity chromatography. The advance in genetic engineering and computational biology approaches leads to development of engineered a nity ligands with improved properties. Computational biology methods such as molecular modeling, molecular docking, molecular dynamic simulations and protein redesign software were applied to design mutated Protein L ligands. The engineered ligands were experimentally studied after being coupled to a solid matrix. The influence of the engineered ligands on the performance of a nity purification with loading prepared Fab were investigated and the dynamic binding capacity, product purity and recovery for the engineered resins were evaluated and compared to Capto™ L resin (GE Healthcare Bio-Sciences AB, Sweden). The highest dynamic binding capacity at ۱۰% breakthrough (DBC۱۰%) was observed for Model-۱ resin; ۱۹.۵ mg/ml, as compared to the DBC۱۰% of Model-۲, Model-۳ and Capto™ L resins; ۷.۲ mg/ml, ۷.۸ mg/mL and ۱۶.۵ mg/mL respectively. Also, recovered Fab fragments from E.coli lysate by Model-۱ and Model-۳ resins was evaluated ۹۵% compared to ۹۲% and ۳۹% by Capto™ L and Model-۲ resins as well as the results of SEC-HPLC analysis showed the purities of recovered Fab fragments by all of resins were over ۹۵%. The results indicated that the Protein L Model-۱ resin was an optimal resin for purification antibody fragments containing kappa light chain from E.coli host cell proteins in a nity chromatography

کلیدواژه ها:

Protein L ، Antibody fragments ، Molecular docking studies ، Molecular dynamic simulations ، OSPREY ، A nity chromatography