Immobilization of an artificial imine reductase within silica nanoparticles improves its performance

Abstract

Silica nanoparticles equipped with an artificial imine reductase display remarkable activity towards cyclic imine- and NAD(+) reduction. The method, based on immobilization and protection of streptavidin on silica nanoparticles, shields the biotinylated metal cofactor against deactivation yielding over 46 000 turnovers in pure samples and 4000 turnovers in crude cellular extracts.

Artificial imine reductases: developments and future directions

“…49,60 Immobilisation of artIR-EDs has provided additional ways to improve stability and recovery strategies. 12,55 In summary, artIREDs have exciting potential biomedical applications in addition to biocatalysis opportunities, in particular their use in enzyme cascades and in whole cell in vivo catalysis. Successes in altering and improving the activity of artIREDs will play a crucial role in designing and developing ArMs, not only for imine reduction, but also other synthetically challenging reactions.…”

“…The nanoparticles could be recycled up to three times with only a minor loss of selectivity. 12 Another study encapsulated S112A, S112K and S112A-K121A Sav artIRED variants inside ferritin, an iron storage protein, which provides a tertiary coordination shell for the solvent exposed catalyst. Encapsulation of the artIRED was achieved by exposing ferritin to low pH, at which the protein would unfold, before raising the pH together with the addition of the artIRED, allowing ferritin to refold around the artIRED.…”

“…Several IREDs have, in addition, been reported to display product inhibition, 11 a common problem of natural enzymes, which can be solved by feeding strategies, but this requires more specialised experimental set-ups. Other concerns, which include a still limited substrate scope, 5 poor long-term stability, 12 high cost of development and the need for expensive cofactors, such as NADH/NADPH [12][13][14] suggest that complementary approaches to natural metalloenzyme discovery are merited for imine reduction.…”

Exploring and Adapting the Molecular Selectivity of Artificial Metalloenzymes

“…For iridium-based ArMs, the majority of literature studies thus far have extensively focused on their use as artificial transfer hydrogenases (ATHase). [113][114][115][116][117][118][119][120][121][122][123][124][125][126][127][128][129][130][131][132] In these works, asymmetric transfer hydrogenation has been shown possible for a number of substrates that include imines, enones, ketones, and for NAD + reduction. One intriguing example of an ATHase that has been used with biological systems relies on a cell-surface display system that shuttles streptavidin into the E. coli periplasm ( Figure 5A).…”

Imine Reductase Based All-Enzyme Hydrogel with Intrinsic Cofactor Regeneration for Flow Biocatalysis

“…For industrial purposes it is convenient to work with cell-free or immobilized enzymes to avoid cross-reactivities and, especially in the case of cofactor dependent enzymes, to gain control over economically attractive regeneration systems. However, only few examples of IREDs immobilized on carrier materials have been reported so far and none have been used for continuous flow biocatalysis [22,23].…”