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Volume 7, Issue 1 (Suppl) J Biotechnol Biomater
ISSN: 2155-952X JBTBM, an open access journal
March 20-21, 2017 Rome, Italy
&
15th World Congress on
2nd International Conference on
Biotechnology And Biotech Industries Meet
Enzymology and Molecular Biology
Enzymology & Mol. Biology 2017
Biotechnology Congress 2017
March 20-21, 2017
Foteini M Pouliou et al., J Biotechnol Biomater 2017, 7:1(Suppl)
Engineering of Tau class GSTs for the development of biosensor
Foteini M Pouliou and Nikolaos E Labrou
Agricultural University of Athens, Greece
Glutathione transferases (GSTs, EC 2.5.1.18) constitute one of the most important families of detoxifying enzymes in nature
with multiple biotechnological applications. GSTs are involved in the detoxification mechanism of endogenous and xenobiotic
electrophile compounds by catalyzing the nucleophilic attack of reduced glutathione (GSH) on the electrophilic center of xenobiotic
compounds including pesticides. This catalytic activity is the basis for the development of enzyme biosensor for herbicide
determination in environmental samples. A library of Tau class GSTs was constructed by DNA shuffling using the DNA encoding
the Glycine max GSTs GmGSTU2-2, GmGSTU4-4 and GmGSTU10-10. The DNA library contained chimeric structures of alternated
segments of the parental sequences and point mutations. Chimeric GST sequences were expressed in Escherichia coli, purified by
affinity chromatography and their enzymatic activities towards CDNB (1-chloro-2,4-dinitrobenzene) were determined. A selected
chimeric enzyme which exhibited high catalytic activity and stability was used for the development of enzyme biosensor. The
inhibition potency of 47 different pesticides towards the chimeric enzyme was evaluated using activity assays. Five compounds, one
insecticide and four fungicides, showed high inhibition potency (IC50) towards the chimeric GST. Kinetic inhibition studies revealed
that pesticides appeared to bind at the substrate-binding region in a competitive manner with respect to the substrate. The chimeric
enzyme will be immobilized and will be explored for the construction of an optical biosensor. This biosensor will be portable, easy to
use, allowing the direct determination of pesticides in environmental samples.
Biography
Foteini M Pouliou is a PhD candidate at the Agricultural University of Athens since 2014. She majored in Biotechnology from the Agricultural University of Athens
in 2012. She has done her Master of Science studies in 2013 focusing on the Bioactive Products and Protein Technology. Her research interests include protein
engineering, enzyme and environmental biotechnology.