The co-immobilization of P450-type nitric oxide reductase and glucose dehydrogenase for the continuous reduction of nitric oxide via cofactor recycling

Garny, Seike; Beeton-Kempen, Natasha; Gerber, Isak B.; Verschoor, J.A.; Jordaan, Justin

doi:10.1016/j.enzmictec.2015.10.006

Cited by 12 publications

(8 citation statements)

References 108 publications

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“…SWV and EIS assays performed using Fe(CN)6 3−/4− as electroactive indicator in the supporting electrolyte (pH = 6.0) were used to characterize the different steps involved in the biosensor development. pH has a marked effect on NOR activity and it was maintained at the optimum value of 6.0 during all the experiments since, at this value, maximum enzyme catalytic activity was detected with the observed protonation of the residues surrounding the catalytic centre [15,39]. pH is a critical point on the development of enzymatic biosensors because it can promote changes in the shape of enzymes and in their Gibbs energy, as well in the ionic charge of the substrate [40].…”

Section: Electrochemical Characterizationmentioning

confidence: 99%

Biosensor for direct bioelectrocatalysis detection of nitric oxide using nitric oxide reductase incorporated in carboxylated single-walled carbon nanotubes/lipidic 3 bilayer nanocomposite

Gomes

Maia

Loureiro

et al. 2019

Bioelectrochemistry

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An enzymatic biosensor based on nitric oxide reductase (NOR; purified from Marinobacter hydrocarbonoclasticus) was developed for nitric oxide (NO) detection. The biosensor was prepared by deposition onto a pyrolytic graphite electrode (PGE) of a nanocomposite constituted by carboxylated single-walled carbon nanotubes (SWCNTs), a lipidic bilayer [1,2-di-(9Z-octadecenoyl)-sn-glycero-3-phosphoethanolamine (DOPE), 1,2-di-(9Z-octadecenoyl)-3-trimethylammonium-propane (DOTAP), 1,2-distearoyl-snglycero-3-phosphoethanolamine-polyethylene glycol (DSPE-PEG)] and NOR. NOR direct electron transfer and NO bioelectrocatalysis were characterized by several electrochemical techniques. The biosensor development was also followed by scanning electron microscopy and Fourier transform infrared spectroscopy. Improved enzyme stability and electron transfer (1.96 × 10 −4 cm.s −1 apparent rate constant) was obtained with the optimum SWCNTs/(DOPE:DOTAP:DSPE-PEG)/NOR) ratio of 4/2.5/4 (v/v/v), which biomimicked the NOR environment. The PGE/[SWCNTs/(DOPE:DOTAP:DSPE-PEG)/NOR] biosensor exhibited a low Michaelis-Menten constant (4.3 μM), wide linear range (0.44-9.09 μM), low detection limit (0.13 μM), high repeatability (4.1% RSD), reproducibility (7.0% RSD), and stability (ca. 5 weeks). Selectivity tests towards Larginine, ascorbic acid, sodium nitrate, sodium nitrite and glucose showed that these compounds did not significantly interfere in NO biosensing (91.0 ± 9.3%-98.4 ± 5.3% recoveries). The proposed biosensor, by incorporating the benefits of biomimetic features of the phospholipid bilayer with SWCNT's inherent properties and NOR bioelectrocatalytic activity and selectivity, is a promising tool for NO.

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Section: Electrochemical Characterizationmentioning

confidence: 99%

Biosensor for direct bioelectrocatalysis detection of nitric oxide using nitric oxide reductase incorporated in carboxylated single-walled carbon nanotubes/lipidic 3 bilayer nanocomposite

Gomes

Maia

Loureiro

et al. 2019

Bioelectrochemistry

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“…Such a strategy might overthrow effects deriving from multiple interactions and also mimic "natural" cascade reactions in which enzymes are usually in close proximity to one another [29]. Co-immobilization is frequently used for co-factor-depending enzymes (i.e., oxidoreductases) to fix on the same carrier both the enzyme catalyzing the main biotransformation and an ancillary enzyme responsible for the regeneration of the co-factor [30][31][32][33]. However, due to the complexity of this approach for our case-study, we here opted to have the enzymes immobilized on the same carrier, but separately, for individual optimization, taking advantage from the accumulated data on CpUP, AhPNP and DddAK in the single bioconversions (phosphorolysis, transglycosylation, and phosphorylation) ( Figure S1).…”

Section: Immobilization Carriermentioning

confidence: 99%

A Multi-Enzymatic Cascade Reaction for the Synthesis of Vidarabine 5′-Monophosphate

et al. 2020

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We here described a three-step multi-enzymatic reaction for the one-pot synthesis of vidarabine 5′-monophosphate (araA-MP), an antiviral drug, using arabinosyluracil (araU), adenine (Ade), and adenosine triphosphate (ATP) as precursors. To this aim, three enzymes involved in the biosynthesis of nucleosides and nucleotides were used in a cascade mode after immobilization: uridine phosphorylase from Clostridium perfringens (CpUP), a purine nucleoside phosphorylase from Aeromonas hydrophila (AhPNP), and deoxyadenosine kinase from Dictyostelium discoideum (DddAK). Specifically, CpUP catalyzes the phosphorolysis of araU thus generating uracil and α-d-arabinose-1-phosphate. AhPNP catalyzes the coupling between this latter compound and Ade to form araA (vidarabine). This nucleoside becomes the substrate of DddAK, which produces the 5′-mononucleotide counterpart (araA-MP) using ATP as the phosphate donor. Reaction conditions (i.e., medium, temperature, immobilization carriers) and biocatalyst stability have been balanced to achieve the highest conversion of vidarabine 5′-monophosphate (≥95.5%). The combination of the nucleoside phosphorylases twosome with deoxyadenosine kinase in a one-pot cascade allowed (i) a complete shift in the equilibrium-controlled synthesis of the nucleoside towards the product formation; and (ii) to overcome the solubility constraints of araA in aqueous medium, thus providing a new route to the highly productive synthesis of araA-MP.

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“…On the other hand, cofactor and enzymes can be co‐immobilized to fabricate self‐sufficient heterogeneous biocatalysts that do not require exogenous supply of cofactor. Even though the concept of self‐sufficient heterogeneous biocatalyst has been exploited for a dozen of cascade reactions, the cofactor utilization inside the solid particles has been rarely studied. Velasco‐Lozano et al .…”

Section: Single‐particle Reaction Kinetics Of Immobilized Enzymesmentioning

confidence: 99%

“…On the other hand, cofactor and enzymes can be co-immobilized to fabricate self-sufficient heterogeneousb iocatalysts that do not require exogenous supply of cofactor.E ven thought he concepto fs elf-sufficient heterogeneous biocatalyst has been exploited for ad ozen of cascade reactions, [40,[71][72][73][74] the cofactoru tilization inside the solid particles has been rarely studied. Velasco-Lozano et al [69] have mapped the cofactor utilization within porous agarose particlest hat co-immobilize the main enzyme (alcohol dehydrogenase), the cofactor recycling enzyme (formate dehydrogenase) and the cofactor (NAD + ).…”

Section: Single-particle Reaction Kinetics Of Immobilized Enzymesmentioning

confidence: 99%

Single‐Particle Studies to Advance the Characterization of Heterogeneous Biocatalysts

et al. 2018

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Immobilized enzymes have been widely exploited because they work as heterogeneous biocatalysts, allowing their recovery and reutilization and easing the downstream processing once the chemical reactions are completed. Unfortunately, we suffer a lack of analytical methods to characterize those heterogeneous biocatalysts at microscopic and molecular levels with spatio‐temporal resolution, which limits their design and optimization. Single‐particle studies are vital to optimize the performance of immobilized enzymes in micro/nanoscopic environments. In this Concept article, we review different analytical techniques that address single‐particle studies to image the spatial distribution of the enzymes across the solid surfaces, the sub‐particle substrate diffusion, the structural integrity and mobility of the immobilized enzymes inside the solid particles, and the pH and O2 internal gradients. From our view, such sub‐particle information elicited from single‐particle analysis is paramount for the design and fabrication of optimal heterogeneous biocatalyst.

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The co-immobilization of P450-type nitric oxide reductase and glucose dehydrogenase for the continuous reduction of nitric oxide via cofactor recycling

Cited by 12 publications

References 108 publications

Biosensor for direct bioelectrocatalysis detection of nitric oxide using nitric oxide reductase incorporated in carboxylated single-walled carbon nanotubes/lipidic 3 bilayer nanocomposite

Biosensor for direct bioelectrocatalysis detection of nitric oxide using nitric oxide reductase incorporated in carboxylated single-walled carbon nanotubes/lipidic 3 bilayer nanocomposite

A Multi-Enzymatic Cascade Reaction for the Synthesis of Vidarabine 5′-Monophosphate

Single‐Particle Studies to Advance the Characterization of Heterogeneous Biocatalysts

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