We further expanded the molecular enzymology study by constructing the in vitro mutagenized A and B transferase constructs that possessed any one of the 20 amino acids at codon 268 (Yamamoto and McNeill, 1996). A transferase possessed the smallest amino acid residue G at codon 268 and expressed only A antigens. When it was replaced with A, the second smallest amino acid residue, the construct expressed small amounts of B antigens in addition to A antigens. When it was replaced with the slightly larger S or cysteine (C), the constructs also expressed smaller amounts of A antigens and larger amounts of B antigens. The N and threonine (T) constructs only expressed small amounts of B antigens. The remaining constructs did not express either A or B antigens with the exception of the histidine (H) construct, which expressed somewhat moderate amounts of A antigens.