Human RBCs blood group conversion from A to O using a novel α-N-acetylgalactosaminidase of high specific activity

α-N-acetylgalactosaminidase (αNAGA) can convert group A human red blood cells (RBCs) to group O. One novel αNAGA gene was cloned by PCR from Elizabethkingia meningosepticum isolated from a domestic clinical sample. Pure recombinant αNAGA was obtained by genetic engineering and protein purification with a calculated molecule of 49.6 kD. αNAGA was selective for terminal α-N-acetylgalacto-samine residue with a high specific activity. αNAGA could completely remove A antigens of 1 U (about 100 mL) group A1 or A2 RBCs in 1 h at pH 6.8 and 25℃ with a consumption of 1.5 or 0.4 mg recombinant enzyme. Enzyme-converted group A RBCs did not agglutinate after being mixed with monoclonal anti-A or sera of groups A,B,AB and O. Other blood group antigens except ABO had no change. FCM analy-sis showed that A antigens and A1 antigens disappeared while H antigens increased. It indicated that αNAGA successfully converted human blood group A RBCs to universally transfusable group O RBCs without the risk of ABO-incompatible transfusion reactions. This αNAGA was suitable for producing universal RBCs to increase clinical transfusion safety,improve the RBCs supply,and to decrease transfusion cost and support transfusion service in case of emergency.

α-N-acetylgalactosaminidase （αNAGA） can convert group A human red blood cells （RBCs） to group O. One novel αNAGA gene was cloned by PCR from Elizabethkingia meningosepticum Isolated from a domestic clinical sample. Pure recombinant αNAGA was obtained by genetic engineering and protein purification with a calculated molecule of 49.6 kD. αNAGA was selective for terminal α-N-acetylgalacto- samine residue with a high specific activity, αNAGA could completely remove A antigens of 1 U （about 100 mL） group A1 or A2 RBCs in 1 h at pH 6.8 and 25℃ with s consumption of 1.5 or 0.4 mg recombinant enzyme. Enzyme-converted group A RBCs did not agglutinate after being mixed with monoclonal snti-A or sere of groups A, B, AB and O. Other blood group antigens except ABO had no change. FCM analysis showed that A antigens and A1 antigens disappeared while H antigens increased. It indicated that αNAGA successfully converted human blood group A RBCs to universally transfusable group O RBCs without the risk of ABO-incompaUble transfusion reactions. This αNAGA was suitable for producing universal RBCs to increase clinical transfusion safety, improve the RBCs supply, and to decrease transfusion cost and support transfusion service in case of emergency.