Amino acid substitutions specific for allotypic G3m(g5) marker were studied by sequence analysis of the C-terminal BrCN peptides of myeloma protein Ba [G3m(g5+)] and Bu [G3m(g5-)]. The results indicate that arginine and tyrosine at position 435 and 436 are responsible for the specificity. Two substitutions of IgG3 Bu [G3m(b1, b3)] and IgG3 Jir [G3m(b3,s,t)] [Matsumoto et al.: J. Immun. 131: 1865, 1983], in the same positions are arginine-phenylalanine and histidine-tyrosine, respectively. Affinity chromatographic data of modified IgG3 proteins show that the tyrosine residue at position 436 associated with phenylalanine at position 124 of protein A-B fragment plays a role in the interaction. The differences in the yields between IgG3s carrying various haplotypes on Protein A-Sepharose affinity chromatography [Ito et al.: Proc. Japan Acad. 56B: 226, 1980] are also explained through this chromatography and also by the configuration of the residues in tertiary structure [Deisenhofer: Biochemistry 20: 2361, 1980].