Mechanism-based catalytic antibody inactivation.
Studies of alternative substrates of the catalytic monoclonal antibodies 18H4, 7D4, and 45A11 provided us with a better understanding of the mechanism of ester hydrolysis employed by these isoabzymes. The antibodies were studied with analogs of the substrate, phenyl acetate; N-acetylglycine phenyl ester 3 and N-carbobenzoxy-glycine phenyl ester 4. All three antibodies catalyzed 3 hydrolysis with kinetic constants similar to those seen with phenyl acetate hydrolysis. However, 4 was found to be a mechanism-based (suicide) inactivator of 18H4 with a kinact of 0.29 min-1 and a K' of 64 microM. Antibody 18H4 was inactivated by 4 after 3.6 turnovers resulting in the acylation of 1.6 tyrosines per combining site. The data conform to a mechanism in which an inactive O-ZGly-tyrosyl-antibody is formed via a Michaelis complex.
Angeles
T S
TS
Department of Enzymology, IGEN, Inc., Rockville, MD 20852.
Martin
M T
MT
D000911:Antibodies, Monoclonal / Q000378:metabolism*; D002384:Catalysis; D004952:Esters; D006867:Hydrolases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D006868:Hydrolysis; D007700:Kinetics; D013379:Substrate Specificity; D013997:Time Factors
D000911:Antibodies, Monoclonal; D004952:Esters; D006867:Hydrolases