Heavy chain variants affect light and heavy chains assembly of monoclonal antibody expressed by Pichia pastoris

Monoclonal Antibody accounts for the largest share of recombinant protein drugs and is the primary choice for the treatment of various diseases. In this study, the monoclonal antibody Eptinezumab was expressed by a yeast host Pichia pastoris. Although the expression and secretion of light chain was efficient, the assembly efficiency between light and heavy chains was low. As the retention of the heavy chain in endoplasmic reticulum may trigger protein degradation, ERAD ubiquitination-related genes were then knocked out separately but it only led to minor improvement effect. Expression of splitted heavy chain variants further revealed that although endoplasmic reticulum retention of the heavy chain upregulated KAR2 expression, it did not affect the assembly efficiency of the light and heavy chains. It was inferred that binding of complete heavy chains to KAR2 spatially affected the assembly between light and heavy chains. Design and screening of KAR2 variants that facilitating full-length antibody assembly could be preferentially considered in future.