Protein A, the gold standard ligand for IgG purification, provides unparalleled removal of host cell proteins (HPCs) and high product capacity. Undesirable qualities of protein A, such as cost, base lability, and low pH elution, have promoted efforts seeking cost-effective alternatives capable of equally efficient separations without the downsides. Synthetic ligands discovered so far enable milder pH elution and excellent reusability, but have not yet managed to reach the 'holy grail' of an LRV &gt; 3. Toward this goal, we utilized known IgG-binding peptides as reference sequences for an <i>in silico</i> search towards variants with improved LRV of CHO HCPs. The sequences with predicted higher affinity were investigated experimentally. Two sequences, V3 and V5, showed excellent values of IgG binding affinity (Kd of 3.22e-6 M and 8.14e-6 M, respectively) and dynamic binding capacities (43.7 mg/mL resin and 55.1 mg/mL resin, respectively, at 5 minute residence time). When evaluated for IgG purification from a CHO cell culture fluids, V3 and V5 afforded a single-pass HCP LRV of 2.7 and 2.3. Further optimization of the ligand density on the chromatographic resin, and the load/wash conditions have enabled further improvements, towards true Protein A-like levels of the IgG purity.

Toward a true Protein A mimetic: new peptide affinity ligand for IgG purification afford high HCP LRV

2019 Spring National Meeting

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