Preparative Production of Acetylcholinesterase and Paraoxonase in Prokaryotic and Eucaryotic Expression Systems

摘要

The acute toxicity of organophosphate (OP) compounds in mammals in general, and in humans in particular, is generally ascribed to their irreversible inhibition of acetyicholinesterase (AChE), the enzyme that terminates the action of acetylcholine (ACh) at cholinergic synapses by its rapid hydrolysis (1). Current medical protection against OPs involves anticholinergic drugs to counteract the accumulation of ACh (2), and quatemary oximes to reactivate OP-inhibited AChE (3). Since some OP compounds, such as soman, inhibit AChE and then rapidly 'age' (4), carbamate pretreatment was developed (5). Although both oximes and carbamylation regimens are partially effective, they have substantial side effects, and do not provide complete protection (6). This led to an interest in non-pharmacological approaches to protection, such as administration of enzyme scavengers (7), or OP hydrolases (8). The feasibility of using bioscavengers, such as AChE or BChE, has been demonstrated in rodents as well as in non-human primates (7). Animal trials have shown that injection of purified human serum paraoxonase (PON1) can protect against OP toxicity (9), and PONi knockout mice display heightened sensitivity to chlorpyrifos oxon relative to wild-type mice (9,10). The hydrolytic capacity of PON1 might render it even more effective relative to the stoichiometric capacity of AChE and BChE. Since wild-type PON does not display high turnover in hydrolysing nerve agents (11), site-directed mutagenesis might also be used to enhance its catalytic activity (12).