title: Proteolytic fragmentation with high specificity of mouse immunoglobulin G. Mapping of proteolytic cleavage sites in the hinge region.
abstract: We report the results of fragmentation of mouse IgG by clostripain, lysyl endopeptidase, metalloendopeptidase, and V8 protease that have a narrower substrate specificity than papain and pepsin. A panel of mouse monoclonal switch variant antibodies with IgG1, IgG2a, and IgG2b subclasses were examined. Cleavage sites by these proteases were mapped on the hinge region of each of the IgG subclasses. It was demonstrated that lysyl endopeptidase can cleave the core hinge portion of IgG2a and IgG2b without perturbing the inter-chain disulfide bridges. Digestion products were successfully isolated by a combined use of protein A affinity chromatography and HPLC techniques. This is a first successful attempt of obtaining the F(ab')2 fragment of IgG2b by proteolytic digestion.
authors.affiliation: Faculty of Pharmaceutical Sciences, University of Tokyo, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Yamaguchi
authors.forename: H
authors.initials: H
authors.lastname: Kim
authors.forename: K
authors.initials: K
authors.lastname: Kato
authors.forename: K
authors.initials: K
authors.lastname: Masuda
authors.forename: I
authors.initials: I
authors.lastname: Shimada
authors.forename: Y
authors.initials: Y
authors.lastname: Arata
mesh_terms: D000478:Alkylation; D000595:Amino Acid Sequence; D000818:Animals; D003546:Cysteine Endopeptidases; D007128:Immunoglobulin Fragments / Q000737:chemistry*; D007074:Immunoglobulin G / Q000737:chemistry*; D008666:Metalloendopeptidases; D051379:Mice; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010084:Oxidation-Reduction; D010206:Papain; D010449:Peptide Mapping; D012697:Serine Endopeptidases
chemical_list: D007128:Immunoglobulin Fragments; D007074:Immunoglobulin G; D012697:Serine Endopeptidases; C028344:glutamyl endopeptidase; C036813:lysyl endopeptidase; D003546:Cysteine Endopeptidases; D010206:Papain; C019913:clostripain; D008666:Metalloendopeptidases