Characterization of upFc, a fragment of human immunoglobulin G1 produced by pepsin in urea.
The digestion of human IgG1/K myeloma proteins with pepsin in the presence of 8 M-urea produces fragments that differ from those produced by aqueous peptic digestion, and from other characteristic immunoglobulin fragments. Fb'2, the larger urea/pepsin fragment, was previously shown to consist of the constant regions of the light chains, and the CH1 domains and hinge regions of the heavy chains. The smaller fragment, upFc, has now been characterized. After reduction, three peptides were released from fragment upFc. Amino acid sequencing, N- and C-terminal determinations and amino acid compositions have enabled these peptides to be identified as residues Ile-253 to Leu-306, residues Thr-307 to Asp-376 and residues Thr-411 to Gly-446 of the heavy chain. Fragment upFc therefore contains the entire Fc region, beginning at residue Ile-253, except for a 34-residue section from within the CH3-domain disulphide loop. Peptic digestion of IgG1/K proteins in 8M-urea therefore provides a method for isolating from gamma1 heavy chains five homogeneous peptides in good yield, which account for almost the entire constant region. Characterization of fragments Fb'2 and upFc has shown that the action of pepsin in urea is entirely different from that of aqueous pepsin. Two gamma1 heavy chains have been shown to differ in sequence at three positions from the sequence reported for protein Eu.
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D006801:Humans; D007141:Immunoglobulin Fc Fragments / Q000032:analysis*; D007074:Immunoglobulin G / Q000032:analysis*; D009194:Myeloma Proteins / Q000032:analysis; D010434:Pepsin A; D010446:Peptide Fragments / Q000302:isolation & purification; D014508:Urea
D000596:Amino Acids; D007141:Immunoglobulin Fc Fragments; D007074:Immunoglobulin G; D009194:Myeloma Proteins; D010446:Peptide Fragments; D014508:Urea; D010434:Pepsin A