Catalytic antibodies induced by a zwitterionic hapten.
A zwitterionic hapten 4 featuring both positively and negatively charged functional groups was designed and synthesized with the goal of generating catalytic antibodies for the hydrolysis of ester 6 and amide 7. Of the 36 monoclonal antibodies specific to BSA-4 (bovine serum albumin) that were isolated, six accelerated the hydrolysis of 6. Two catalytic antibodies with distinctively different and representative kinetic behaviors were selected for detailed kinetic studies. Whereas H8-2-6F11 showed burst kinetic behavior, which can be attributed to the formation of an acyl intermediate, H8-1-2D5 did not, but it did exhibit high multiple turnover activity. The rate of hydrolysis of 6 catalyzed by H8-1-2D5 followed Michaelis-Menten kinetics; the apparent values of the Michaelis-Menten constant Km and the catalytic constant kcat were 488 microM and 3.5 min(-1), respectively. The catalytic rate enhancement (kcat/kun) observed for H8-1-2D5 was 1.3 x 10(5), which is approximately two orders of magnitude greater than those for monofunctional haptens. Thus H8-1-2D5 compares well in catalytic activity with antibodies isolated by a related approach called heterologous immunization.
Tsumuraya
T
T
Kao Institute for Fundamental Research, Ichikaimachi, Haga, Tochigi, Japan. tsumu@beri.co.jp
Takazawa
N
N
Tsunakawa
A
A
Fleck
R
R
Masamune
S
S
D000577:Amides / Q000378:metabolism; D000818:Animals; D017151:Antibodies, Catalytic / Q000096:biosynthesis / Q000737:chemistry*; D001666:Binding Sites, Antibody; D004952:Esters / Q000378:metabolism; D006241:Haptens / Q000008:administration & dosage / Q000276:immunology*; D006868:Hydrolysis; D007477:Ions; D007700:Kinetics; D051379:Mice; D008956:Models, Chemical; D013329:Structure-Activity Relationship
D000577:Amides; D017151:Antibodies, Catalytic; D004952:Esters; D006241:Haptens; D007477:Ions