Inhibition of -SH enzymes by an impurity in commercial samples of diisopropylphosphofluoridate

Alkylphosphorylation of Stem Bromelain by Diisopropylphosphorofluoridate without Inhibition of Proteinase Activity*

“…The conclusion we have drawn from the results of the earlier and the present experiments is that DFP per se has no inhibitory effect on stem bromelain, while impurities associated with some DFP preparations may cause an inhibition by irreversibly attacking the essential SH group of the enzyme. A similar conclusion has been arrived at more recently by Gould et al (1963) and Gould and Liener (1965) from the experiments with ficin and papain. They have demonstrated that there exists in some commercial preparations of DFP an impurity which reacts with sulfhydryl (SH) groups of cysteine and enzyme proteins.…”

“…The attack by the impurity must be of such a nature that it can be protected by excess cysteine but not by cyanide. Using ficin, Gould et al (1963) and Gould and Liener (1965) have found that there exists in some commercial preparations of DFP an impurity which reacts with SH groups. They fractionated a specimen of DFP from Aldrich Chemical Co., Milwaukee, Wis., by distillation under reduced pressure into four fractions.…”

Enzyme Hydrolysis, Purification and Properties of Enzyme Responsible for Hydrolyzing Phenylcarbamates

“…This implies that a hydroxyl group, probably the hydroxyl group of serine, is involved at the active center (20). Interpretation of DFP data, however, must be viewed with some caution since some commercial preparations are contaminated with a sulfhydryl inhibitor (7). The enzyme under study, however, is not inhibited by />-hydroxymercuribenzoate at concentrations up to 10 ~3M in the assay system; nor is the enzyme enhanced by the addition of reduced glutathione.…”

“…Insight into the factors responsible for altering the rates of hydrolysis of certain substrates can be gained by examining a physical property of the phenylcarbamates. Camper and Moreland (7) have recently studied the relative acidities of many of the same phenylcarbamates used in the study of substrate specificity reported in this paper. It is of interest, then, to determine w-hether any correlation exists between the relative acidities of the phenylcarbamates and their susceptibility to enzymic hydrolysis.…”

“…Relative acidities of the phenylamides are determined by potentiometric titration with 0.1 A' tetrabutvlammonium hydroxide methoxide in «-butylamine (7).…”