Chemistry of Succinimido Esters. XIX

Chemical modification (N-arylacetylation) of amino groups in proteins by N-succinimidyl arylacetates (1a) was studied kinetically using bovine serum albumin (BSA) as the model protein in the range of 7.0≤pH≤9.0 at 20°C. N-Arylacetylation was competitive with the hydrolysis of (1a). Its rate was expressed as V= k2 [NH2 of BSA] [1a].The logarithm of the second order rate constant, k2, increased linearly with the relative acid strength of the corresponding benzoic acid derivatives, log (K/K0), for meta-substituted (1a). The slope was independent of pH. The rate of ortho-substituted (1a) was less than that expected from log (K/ K0) owing to steric hindrance of the substituent. pH profiles gave slopes of less than unity since the amino groups of BSA were not equivalent. The rate ratio, 10-3 k2/k1, where k1 is the first order rate constant for the hydrolysis of (1a), exceeded unity for all cases and was maximal at about pH 8.5. The reactivity of (1a) was compared with those of N-succinimidyl benzoates (1b) and N-acyloxysuccinimides (1c).Strong hydrophobic interaction between BSA and the imido ester was observed for acylation by (1c) with a long carbon chain. However, little or no significant rate acceleration could be observed for (1a) (except p-chlorosubstituent) or (1b) because of the weaker hydrophobicity of the arylacetyl and aryl groups compared with the long alkyl chain.Based on the present results, (1a) is concluded applicable for use as an N-arylacetylation reagent for the chemical modification of Lys and terminal amino groups in proteins. A possible mechanism is also proposed and discussed.