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Team:Exeter/invivo
From 2014.igem.org
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Measuring in vivo Degradation Using Metabolite Colour Change
Introduction
Aim
The aim of this experiment was to measure the degradation rate of trinitrotoluene (TNT) and nitroglycerin (NG) in vivo using Raman spectroscopy. The degradation rate of enzymes Nem-A and Xen-B when transcribed by an E. coli bacterium will be measured. In addition to the degradation rates, a value for the partition coefficient for TNT and NG between water and the cell membrane was determined.
TNT Degradation
During the NemA and XenB-catalysed degradation of TNT, a series of nitrite groups as well aromatic ring reduction leads to formation of amino-dimethyl-tetranitrobiphenyl. During this process a hydride-Meisenheimer complex metabolite is formed. This metabolite has a distinct dark-brown colour [Vorbeck et.al 1994]. This metabolite causes reaction mixtures with XenB or NemA, mixed with TNT, to change from colourless to red, then to yellow [Pak 2000].
References
- Vorbeck, Claudia; Lenke, Hiltrud; Fischer, Peter; Hans-Joachim, Knackmuss (1994) Identification of a Hydride-MeisenheimerComplex as a Metabolite of 2,4,6-Trinitrotoluene by a Mycobacterium Strain ; Journal of Bacteriology
- Jeong W. Pak; Kyle L. Knoke; Daniel R. Noguera; Brian G. Fox; Glenn H. Chambliss (2000) Transformation of 2,4,6-Trinitrotoluene by Purified Xenobiotic Reductase B from Pseudomonas fluorescens I-C; Applied and Environmental Microbiology
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