(190o) In Situ Growth of Acetylcholinesterase-Oxime Polymer Conjugate Scavengers of Organophosphate Nerve Agent Toxicity

In Situ Growth of Acetylcholinesterase-Oxime Polymer
Conjugate Scavengers of Organophosphate Nerve Agent Toxicity

Libin Zhang^†,
Nicholas Harris^†€, Weihang Ji^†,
Hironobu Murata^†, Krzysztof Matyjaszewski^¡ì,
and Alan J. Russell^†,‡,

^†Center for Polymer-based Protein
Engineering, ICES, Carnegie Mellon University, 5000
Forbes Avenue, Pittsburgh, Pennsylvania 15213, United States

^‡Department of Biomedical
Engineering, Scott Hall 4N201, Carnegie Mellon University, 5000 Forbes Avenue,
Pittsburgh, Pennsylvania 15213, United States

^¡ìDepartment of Chemistry, Carnegie
Mellon University, 4400 Fifth Avenue, Pittsburgh, Pennsylvania 15213, United
States

^€Department of Biotechnology
Engineering, ORT Braude Academic College, Karmiel POB78, Israel

Abstract

Acetylcholinesterase
(AChE) is a serine hydrolase which rapidly catalyzes hydrolysis of the neurotransmitter,
acetylcholine (ACh, Figure 1A). In keeping with its
activity, AChE serves to terminate synaptic
transmission in neuromuscular junctions and maintain a tolerable level of
diffusing substrate, ACh. Organo-phosphorus
(OP) compounds, nerve agents, inhibit AChE causing ACh accumulation in cholinergic synapses. The toxic effects
of accumulated ACh range from excessive salivation
through muscular paralysis to respiratory failure. Nerve agent toxicity can be
treated by intravenous injection of the OP scavenger, butyrylcholinesterase
(BChE).¹ Strong nucleophiles, such as
oximes, are used for reactivation of OP-AChE
conjugates by displacing the OP moiety from the enzyme's active site (Figure
1B) and deactivating circulating Ops. Quaternary pyridinium
oximes, for example, are used for nucleophilic reactivation of AChE that is covalently inhibited by OPs.² Treatment by pyridinium
oximes, however, requires rapid administration after exposure to OPs and is
limited by a short half-life in the blood.  Attachment of synthetic
polymers to form protein-polymer conjugates combines properties derived from
both biologic and synthetic materials. Such conjugates have become central
components of the biologic drug and food industries. Atom transfer radical
polymerization (ATRP), that was developed in the early 1990s, has proven to be
a powerful technique with control of polymer chain length and molecular weight,
producing a narrow size distribution and a large library of available
architectures.³ We have previously generated a series of
protein-polymer conjugates using ATRP to increase protein pH and temperature
stability, enable dissolution of protein in organic solvent and enhance
trans-epithelial protein transport.^4-6 We will describe the design,
synthesis and initial functional characterization of AChE-polymer
conjugates, with oxime pendant on the polymer chain, to study self-reactivation
following inhibition.

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