Structural effect of a recombinant monoclonal antibody on hinge region peptide bond hydrolysis.
IgG hinge region peptide bonds are susceptible to degradation by hydrolysis. To study the effect of Fab and Fc on hinge region peptide bond hydrolysis, a recombinant humanized monoclonal IgG1 antibody, its F(ab')2 fragment, and a model peptide with amino acid sequence corresponding to the hinge region were incubated at 40 degrees C in formulation buffer including complete protease inhibitor and EDTA for 0, 2, 4, 6 and 8 weeks. Two major cleavage sites were identified in the hinge region of the intact recombinant humanized monoclonal antibody and its F(ab')2 fragment, but only one major cleavage site of the model peptide was identified. Hinge region peptide bond hydrolysis of the intact antibody and its F(ab')2 fragment degraded at comparable rates, while the model peptide degraded much faster. It was concluded that Fab region of the IgG, but not Fc portion had significant effect on preventing peptide bond cleavage by direct hydrolysis. Hydrolysis of hinge region peptide bonds was accelerated under both acidic and basic conditions.
Xiang
Tao
T
Process Sciences Department, Abbott Bioresearch Center, 100 Research Drive, Worcester, MA 01605, USA.
Lundell
Edwin
E
Sun
Zuping
Z
Liu
Hongcheng
H
D000911:Antibodies, Monoclonal / Q000737:chemistry*; D004492:Edetic Acid / Q000737:chemistry; D006801:Humans; D006868:Hydrolysis; D007140:Immunoglobulin Fab Fragments / Q000737:chemistry*; D007074:Immunoglobulin G / Q000737:chemistry; D013058:Mass Spectrometry; D008954:Models, Biological; D010455:Peptides / Q000737:chemistry*; D011994:Recombinant Proteins / Q000737:chemistry*; D013997:Time Factors
D000911:Antibodies, Monoclonal; D007140:Immunoglobulin Fab Fragments; D007074:Immunoglobulin G; D010455:Peptides; D011994:Recombinant Proteins; D004492:Edetic Acid