C ‐terminal Lys or Arg Clipping in Proteins

Many secreted proteins are often truncated in their C- and/or N-termini. Before and after secretion, proteins may come in contact with proteases that might specifically clip C- or N-terminal amino acid residues. For example, carboxypeptidases cleaves C-terminal Lys or Arg residues from proteins. Since the enzymatic reaction of carboxypeptidases is often incomplete, such an enzymatic reaction may generate additional heterogeneity of proteins. C-terminal clipping of Lys and Arg residues is very commonly observed in monoclonal antibodies (mAbs) produced using mammalian cells such as Chinese Hamster Ovary (CHO) cells that contain secreted carboxypeptidases in the cell culture media. C-terminal heterogeneity is often analyzed and quantitated using ion-exchange chromatography (IEC), hydrophobic interaction chromatography (HIC), iso-electric focusing (IEF), capillary iso-electric focusing (cIEF), etc., and also by high-resolution mass spectrometry. Although C-terminal heterogeneity may not impact the antibody binding to antigen, it has been recently reported that C-terminal Lys clipping may impact the complement-dependent cytotoxicity (CDC) activity of mAbs.