Cold pepsin digestion: a novel method to produce the Fv fragment from human immunoglobulin M.

• DOI: 10.1073/pnas.75.6.2649
• Publication Date: 2006-05-31T07:43:38Z
• Authors: L C Lin; F W Putnam
• Keywords: Cold Temperature; 0301 basic medicine; 03 medical and health sciences; Immunoglobulin M; Hydrolysis; Immunoglobulin Variable Region; Humans; Binding Sites, Antibody; Immunoglobulin Fragments; Pepsin A; Substrate Specificity; 3. Good health

A strategy for the proteolytic fragmentation of human IgM has been developed. This method is called "cold pepsin digestion" because of its unique feature of achieving restricted peptic cleavages at 4 degrees and pH 4.0. Cold pepsin digestion has been applied successfully to produce an Fv fragment from 14 human IgM proteins. The Fv fragment consists of the heavy chain variable domain (VH) and the light chain variable domain (VL) held together by strong noncovalent interaction. Thus, each Fv fragment contains one intact antigen-binding site and represents the minimal active fragment derivable from an antibody molecule. A series of other structurally and functionally important fragments were also isolated and characterized. Two basic digestion pathways were recognized; these mainly reflect the relative accessibility of five sets of major interdomain cleavage sites.