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Article Dans Une Revue Biosensors and Bioelectronics Année : 2017

Thiamine biosensor based on oxidative trapping of enzyme-substrate intermediate.

Résumé

In the present work, we describe a new thiamine amperometric biosensor based on thiamine pyrophosphate (ThDP)-dependent transketolase (TK)-catalyzed reaction, followed by the oxidative trapping of TK intermediate α,β-dihydroxyethylthiamine diphosphate (DHEThDP) within the enzymatic active site. For the biosensor design purpose, TK from Escherichia coli (TKec) was immobilized in Mg2Al-NO3 Layered Double Hydroxides (LDH) and the electrochemical detection was achieved with the TKec/LDH modified glassy carbon electrode (GCE). The transduction process was based on the ability of Fe(CN)63− to oxidize DHEThDP to glycolic acid along with ThDP regeneration. The released Fe(CN)64− was re-oxidized at +0.5 V vs Ag-AgCl and the reaction was followed by chronoamperometry. The TKec/LDH/GCE biosensor was optimized using the best TK donor substrates, namely l-erythrulose and d-fructose-6-phosphate. ThDP was assayed with great sensitivity (3831 mA M−1 cm−2) over 20–400 nM linear range.

Domaines

Chimie
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Dates et versions

hal-01546142 , version 1 (23-06-2017)

Identifiants

Citer

Matilte Halma, Bastien Doumèche, Laurence Hecquet, Vanessa Prévot, Christine Mousty, et al.. Thiamine biosensor based on oxidative trapping of enzyme-substrate intermediate. . Biosensors and Bioelectronics, 2017, 87, pp.850-857. ⟨10.1016/j.bios.2016.09.049⟩. ⟨hal-01546142⟩
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