A fast and sensitive colorimetric assay (FRED, fast and reliable ene-reductases detection) that allows the estn. of levels of conversion of ene-reductase (ER)-catalyzed reactions has been developed. The activated olefin is reduced by ER at the expense of NAD(P)H cofactor, whose regeneration is carried out in situ by the glucose/glucose dehydrogenase system. Subsequently, the consumption of the co-substrate glucose is detd. colorimetrically by a multienzymic system. The FRED assay offers a wide range of possible applications, from enzyme fingerprinting and kinetic anal., to primary screening of enzyme libraries and optimization of ERs' performances under different reaction conditions.

A Rapid and High-Throughput Assay for the Estimation of Conversions of Ene-Reductase-Catalysed Reactions

CROTTI, MICHELE;GATTI, FRANCESCO GILBERTO;PARMEGGIANI, FABIO;BRENNA, MARIA ELISABETTA;
2015

Abstract

A fast and sensitive colorimetric assay (FRED, fast and reliable ene-reductases detection) that allows the estn. of levels of conversion of ene-reductase (ER)-catalyzed reactions has been developed. The activated olefin is reduced by ER at the expense of NAD(P)H cofactor, whose regeneration is carried out in situ by the glucose/glucose dehydrogenase system. Subsequently, the consumption of the co-substrate glucose is detd. colorimetrically by a multienzymic system. The FRED assay offers a wide range of possible applications, from enzyme fingerprinting and kinetic anal., to primary screening of enzyme libraries and optimization of ERs' performances under different reaction conditions.
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Utilizza questo identificativo per citare o creare un link a questo documento: https://hdl.handle.net/11311/969817
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