Evolved stereoselective hydrolases for broad-spectrum G-type nerve agent detoxification

Area of Science:

• Biochemistry and Molecular Biology
• Enzyme Engineering
• Chemical Warfare Defense

Background:

• Catalytic scavenging by enzymes is a key strategy for neutralizing toxic nerve agents before they can cause harm.
• Serum paraoxonase 1 (PON1) is an enzyme known for its potential in hydrolyzing certain toxic compounds.

Purpose of the Study:

• To enhance the catalytic activity of a PON1 variant for the hydrolysis of toxic S(P) isomers of G-type nerve agents.
• To improve the stereospecificity of PON1 towards the more toxic isomers of nerve agents.

Main Methods:

• Employed directed evolution techniques to simultaneously enhance enzyme activity and alter substrate specificity.
• Selected for variants that prevent acetylcholinesterase inhibition, a key indicator of nerve agent toxicity.
• Quantified catalytic rates and efficiencies for nerve agent hydrolysis.

Main Results:

• Developed evolved PON1 variants with up to 340-fold increased hydrolysis rates for G-type nerve agents.
• Achieved high catalytic efficiencies ranging from 0.2 to 5 × 10^7 M(-1) min(-1).
• Completely reversed stereospecificity, favoring hydrolysis of the toxic S(P) isomer (E > 2,500) over the R(P) isomer (< 6.3 × 10^-4).

Conclusions:

• The evolved PON1 variants demonstrate significantly enhanced catalytic activity and specificity against G-type nerve agents.
• These engineered enzymes represent a potential breakthrough for developing broad-range prophylactic agents against nerve agent intoxication.