Part:BBa_K216006:Experience

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Applications of BBa_K216006

Edinburgh iGEM team, 2009: pentaerythritol tetranitrate reductase from Enterobacter cloacae PB2 is a member of the Old Yellow Enzyme family of flavoprotein oxidoreductases. It is reduced by NADPH. The reduced form of the enzyme is able to reductively denitrate nitrate esters such as pentaerythritol tetranitrate (PETN or penthrite, a common component of plastic explosives) and nitroglycerine (glycerol trinitrate, GTN) yielding the alcohol and nitrite ion. Nitrate groups are removed much more rapidly from compounds with three or more nitrate groups than from those with two or fewer. PETN reductase also reduces di- and trinitroaromatic compounds such as TNT (2,4,6-trinitrotoluene) initially to a hydride-Meisenheimer complex, then a dihydride adduct, and finally to unidentified non-aromatic compounds, with liberation of nitro-groups as nitrite ion. PETN reductase is also reported to have activity against nitramine explosives such as RDX (Royal Demolition Explosive, cyclonite, hexahydro-1,3,5-trinitro-1,3,5-triazine) and HMX (High Melting Explosive, octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine). It is thus able to attack and detoxify all of the major classes of secondary high explosive, liberating nitrogen as nitrite. It therefore has been applied to two applications: as an element in biosensors for explosive detection, based on detection of liberated nitrite; and for bioremediation of toxic explosive residues in contaminated soil, for example, in the form of transgenic plants expressing PETN reductase (French et al, 1999).

At the time of writing, we have not been able to confirm enzyme activity of this construct as our permit to work with explosives has not yet come through. The best assay is Griess assay for nitrite release from either pentaerythritol tetranitrate (PETN) or nitroglycerine (glycerol trinitrate, GTN): see references for protocol. We hope to carry out these assays as soon as we are able to work with these compounds.

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