Substituent Effects on an Antibody-Catalyzed Hydrolysis of Phenyl Esters: Further Evidence for an Acyl-Antibody Intermediate

Richard A. Gibbs; Patricia A. Benkovic; Kim D. Janda; Richard A. Lerner; Stephen J. Benkovic

doi:10.1021/ja00035a057

Substituent Effects on an Antibody-Catalyzed Hydrolysis of Phenyl Esters: Further Evidence for an Acyl-Antibody Intermediate

Richard A. Gibbs, Patricia A. Benkovic, Kim D. Janda, Richard A. Lerner, Stephen J. Benkovic

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Abstract

The hydrolysis of a series of para-substituted phenyl esters (2 and 6a-d) by the monoclonal catalytic antibody NPN43C9 has been investigated. The apparent pK_a observed in the k_cat/K_m-pH profiles shifts from 8.9 (p-nitrophenyl ester) up to a maximum of 9.5 (p-methylphenyl ester). A correlation of the antibody-catalyzed rates of hydrolysis of the esters versus the σ parameter affords a ρ value of 2.3, indicative of a hydrolytic mechanism proceeding via the attack of a neutral nitrogen nucleophile and contrasting with the low ρ value expected for general base catalyzed hydrolysis. There was an inverse solvent deuterium isotope effect on the antibody-catalyzed hydrolysis of the p-chlorophenyl ester at pH > 10. These observations are consistent with our previous proposal (Benkovic, et al. Science 1990, 250, 1135) that this antibody employs a multistep kinetic pathway which involves the formation of a covalent acyl-antibody intermediate.

Original language	English (US)
Pages (from-to)	3528-3534
Number of pages	7
Journal	Journal of the American Chemical Society
Volume	114
Issue number	9
DOIs	https://doi.org/10.1021/ja00035a057
State	Published - Apr 1 1992

All Science Journal Classification (ASJC) codes

Catalysis
General Chemistry
Biochemistry
Colloid and Surface Chemistry

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title = "Substituent Effects on an Antibody-Catalyzed Hydrolysis of Phenyl Esters: Further Evidence for an Acyl-Antibody Intermediate",

abstract = "The hydrolysis of a series of para-substituted phenyl esters (2 and 6a-d) by the monoclonal catalytic antibody NPN43C9 has been investigated. The apparent pKa observed in the kcat/Km-pH profiles shifts from 8.9 (p-nitrophenyl ester) up to a maximum of 9.5 (p-methylphenyl ester). A correlation of the antibody-catalyzed rates of hydrolysis of the esters versus the σ parameter affords a ρ value of 2.3, indicative of a hydrolytic mechanism proceeding via the attack of a neutral nitrogen nucleophile and contrasting with the low ρ value expected for general base catalyzed hydrolysis. There was an inverse solvent deuterium isotope effect on the antibody-catalyzed hydrolysis of the p-chlorophenyl ester at pH > 10. These observations are consistent with our previous proposal (Benkovic, et al. Science 1990, 250, 1135) that this antibody employs a multistep kinetic pathway which involves the formation of a covalent acyl-antibody intermediate.",

author = "Gibbs, {Richard A.} and Benkovic, {Patricia A.} and Janda, {Kim D.} and Lerner, {Richard A.} and Benkovic, {Stephen J.}",

year = "1992",

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doi = "10.1021/ja00035a057",

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T1 - Substituent Effects on an Antibody-Catalyzed Hydrolysis of Phenyl Esters

T2 - Further Evidence for an Acyl-Antibody Intermediate

AU - Gibbs, Richard A.

AU - Benkovic, Patricia A.

AU - Janda, Kim D.

AU - Lerner, Richard A.

AU - Benkovic, Stephen J.

PY - 1992/4/1

Y1 - 1992/4/1

N2 - The hydrolysis of a series of para-substituted phenyl esters (2 and 6a-d) by the monoclonal catalytic antibody NPN43C9 has been investigated. The apparent pKa observed in the kcat/Km-pH profiles shifts from 8.9 (p-nitrophenyl ester) up to a maximum of 9.5 (p-methylphenyl ester). A correlation of the antibody-catalyzed rates of hydrolysis of the esters versus the σ parameter affords a ρ value of 2.3, indicative of a hydrolytic mechanism proceeding via the attack of a neutral nitrogen nucleophile and contrasting with the low ρ value expected for general base catalyzed hydrolysis. There was an inverse solvent deuterium isotope effect on the antibody-catalyzed hydrolysis of the p-chlorophenyl ester at pH > 10. These observations are consistent with our previous proposal (Benkovic, et al. Science 1990, 250, 1135) that this antibody employs a multistep kinetic pathway which involves the formation of a covalent acyl-antibody intermediate.

AB - The hydrolysis of a series of para-substituted phenyl esters (2 and 6a-d) by the monoclonal catalytic antibody NPN43C9 has been investigated. The apparent pKa observed in the kcat/Km-pH profiles shifts from 8.9 (p-nitrophenyl ester) up to a maximum of 9.5 (p-methylphenyl ester). A correlation of the antibody-catalyzed rates of hydrolysis of the esters versus the σ parameter affords a ρ value of 2.3, indicative of a hydrolytic mechanism proceeding via the attack of a neutral nitrogen nucleophile and contrasting with the low ρ value expected for general base catalyzed hydrolysis. There was an inverse solvent deuterium isotope effect on the antibody-catalyzed hydrolysis of the p-chlorophenyl ester at pH > 10. These observations are consistent with our previous proposal (Benkovic, et al. Science 1990, 250, 1135) that this antibody employs a multistep kinetic pathway which involves the formation of a covalent acyl-antibody intermediate.

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JO - Journal of the American Chemical Society

JF - Journal of the American Chemical Society

IS - 9

ER -

Substituent Effects on an Antibody-Catalyzed Hydrolysis of Phenyl Esters: Further Evidence for an Acyl-Antibody Intermediate

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