Isolation and properties of some molecular species of human brain esterases

Abstract

Preparations of the carboxylic ester hydrolases of human brain were obtained by (NH 4) 2SO 4 fractionation and DEAE-Sephadex chromatogrphy procedures. Organophosphate-sensitive esterases (B-type) were precipitated 40% (NH 4) 2SO 4 saturation. Esterases resistant to organophosphate derivatives (A-type) and activated by p-chloromercuribenzoate (PCMB) (C-type) were precipitated from the remaining supernatant by 65% (NH 4) 2SO 2 saturation. The A- and C-type esterases were further fractionated by column chromatography into cathodic- and slow and fast anodic-migrating enzymes. 1. 1. The major anodic enzymes showed hydrolytic activity toward aryl esters of acetic, propionic and butyric acids and were devoid of peptides, amidase or cholinesterase activity. 2. 2. The faster-migrating anodic enzymes were shown by starch-gel electrophoresis to be inactivated by air oxidation and could be reactivated by reducing agents. Reducing agents also reverse the inactivation of the enzyme by 8 M urea. The C-type esterase remained active only in the presence of sulhydryl-reacting compounds or reducing agents capable of forming sulfhydryl derivatives. The esterases of intermediate mobility are inactivated by high concentration of reducing agents or sulfhydryl-reacting compounds and irreversibly inactivated by 8 M urea. 3. 3. It was concluded that the fast-moving esterases require either free SH groups or SH groups reacted with small reducing molecules, such as iodoacetamide or mercaptol derivatives, in order to maintain the conformation required for activity. In contrast, the intermediary anodic esterases appear to require both free SH and S-S groups for activity, since they are inhibited by PCMB and inactivated by high concentrations of reducing agents. It appears that the 3 esterase groups described are SH-enzymes, but differ from each other in the role of the SH group in the determination of enzyme activity.