Protein film electrochemistry
(PFE) has given unrivalled insight
into the properties of redox proteins and many electron-transferring
enzymes, allowing investigations of otherwise ill-defined or intractable
topics such as unstable Fe–S centers and the catalytic bias
of enzymes. Many enzymes have been established to be reversible electrocatalysts
when attached to an electrode, and further investigations have revealed
how unusual dependences of catalytic rates on electrode potential
have stark similarities with electronics. A special case, the reversible
electrochemistry of a photosynthetic enzyme, ferredoxin-NADP+reductase (FNR), loaded at very high concentrations in the 3D nanopores
of a conducting metal oxide layer, is leading to a new technology
that brings PFE to myriad enzymes of other classes, the activities
of which become controlled by the primary electron exchange. This
extension is possible because FNR-based recycling of NADP(H) can be
coupled to a dehydrogenase, and thence to other enzymes linked in
tandem by the tight channelling of cofactors and intermediates within
the nanopores of the material. The earlier interpretations of catalytic
wave-shapes and various analogies with electronics are thus extended
to initiate a field perhaps aptly named “cascade-tronics”,
in which the flow of reactions along an enzyme cascade is monitored
and controlled through an electrochemical analyzer. Unlike in photosynthesis
where FNR transduces electron transfer and hydride transfer through
the unidirectional recycling of NADPH, the “electrochemical
leaf” (e-Leaf) can be used to drive reactions in both oxidizing
and reducing directions. The e-Leaf offers a natural way to study
how enzymes are affected by nanoconfinement and crowding, mimicking
the physical conditions under which enzyme cascades operate in living
cells. The reactions of the trapped enzymes, often at very high local
concentration, are thus studied electrochemically, exploiting the
potential domain to control rates and direction and the current–rate
analogy to derive kinetic data. Localized NADP(H) recycling is very
efficient, resulting in very high cofactor turnover numbers and new
opportunities for controlling and exploiting biocatalysis.