In situ Regeneration of NADH via Lipoamide Dehydrogenase-catalyzed Electron Transfer Reaction Evidenced by Spectroelectrochemistry

Journal Article · Wed Aug 01 04:00:00 EDT 2012 · Bioelectrochemistry and Bioenergetics

DOI:https://doi.org/10.1016/j.bioelechem.2012.03.002· OSTI ID:1051189

Tam, Tsz Kin; Chen, Baowei; Lei, Chenghong; Liu, Jun

NAD/NADH is a coenzyme found in all living cells, carrying electrons from one reaction to another. We report on characterizations of in situ regeneration of NADH via lipoamide dehydrogenase (LD)-catalyzed electron transfer reaction to regenerate NADH using UV-vis spectroelectrochemistry. The Michaelis-Menten constant (Km) and maximum velocity (Vmax) of NADH regeneration were measured as 0.80 {+-} 0.15 mM and 1.91 {+-} 0.09 {micro}M s-1 in a 1-mm thin-layer spectroelectrochemical cell using gold gauze as the working electrode at the applied potential -0.75 V (vs. Ag/AgCl). The electrocatalytic reduction of the NAD system was further coupled with the enzymatic conversion of pyruvate to lactate by lactate dehydrogenase to examine the coenzymatic activity of the regenerated NADH. Although the reproducible electrocatalytic reduction of NAD into NADH is known to be difficult compared to the electrocatalytic oxidation of NADH, our spectroelectrochemical results indicate that the in situ regeneration of NADH via LD-catalyzed electron transfer reaction is fast and sustainable and can be potentially applied to many NAD/NADH-dependent enzyme systems.

• Research Organization: Pacific Northwest National Laboratory (PNNL), Richland, WA (US)
• Sponsoring Organization: USDOE
• DOE Contract Number: AC05-76RL01830
• OSTI ID: 1051189
• Report Number(s): PNNL-SA-83118; 400412000
• Journal Information: Bioelectrochemistry and Bioenergetics, Journal Name: Bioelectrochemistry and Bioenergetics Vol. 86; ISSN BEBEBP; ISSN 0302-4598
• Country of Publication: United States
• Language: English

Related Subjects

59 BASIC BIOLOGICAL SCIENCES
60 APPLIED LIFE SCIENCES
Biocatalytic
COENZYMES
ELECTRODES
ELECTRON TRANSFER
ELECTRONS
ENZYMES
Electron
GOLD
LACTATE DEHYDROGENASE
LACTATES
NADH
OXIDATION
OXIDOREDUCTASES
REGENERATION
Spectroelectrochemistry
VELOCITY
acid
cyclic
electrocatalytic
electrochemical
electrode
electrodes
enzyme
film-modified
glassy-carbon
mediator
methyl
nad(+)
nanotubes
reduction
regeneration
transfer
viologen
voltammetry