Retuning the potential of the electrochemical leaf

Area of Science:

• Biocatalysis
• Electrochemistry
• Enzyme Engineering

Background:

• The electrochemical leaf utilizes electrified enzymes for multi-enzyme cascades.
• Ferredoxin NADP+ reductase (FNR) is key for NADP+/NADPH interconversion within electrodes.
• Co-entrapment of enzymes allows electrical control of extended cascades via NADP(H) recycling.

Purpose of the Study:

• To electrochemically study a variant FNR with an active-site tyrosine-to-serine mutation.
• To investigate the impact of this mutation on cofactor preference and electrochemical properties.
• To understand the catalytic behavior and limitations of the variant FNR.

Main Methods:

• Electrochemical characterization of the variant FNR.
• Enzyme activity assays monitoring NAD(H) turnover.
• Potential-dependent analysis of enzyme kinetics and inhibition.
• Investigation of trapped intermediate states.

Main Results:

• The variant FNR exhibits inverted cofactor preference (NAD(H) over NADP(H)).
• The mutation alters the flavin reduction potential, making it less reductive.
• A trapped intermediate state was observed, requiring negative overpotential for relief.
• NADP+ inhibition of NAD(H) turnover was identified and found to be potential-dependent.

Conclusions:

• The modified FNR offers new possibilities for bioelectrocatalysis by altering cofactor specificity.
• Understanding the electrochemical behavior and intermediate states is crucial for optimizing enzyme cascades.
• These findings advance the development and application of the electrochemical leaf technology.