Event Title

Glucose Oxidase as an Oxygen Scrubber

Mentor 1

Andy Pacheco

Location

Union Wisconsin Room

Start Date

24-4-2015 10:30 AM

End Date

24-4-2015 11:45 AM

Description

Cytochrome C Nitrate Reductase is an anerobic enzyme that shows promise for study to generate raw material for fertilizers. Since the enzyme requires oxygen-free envrionments, hyper-anerobic environments were sought in an anerobic glove box using glucose oxidase to remove trace oxygen. Glucose oxidase catalyses the oxidation of glucose by oxygen to yield gluconolactone and hydrogen peroxide via a ping-pong bi-bi mechanism. The glucose/glucose oxidase pair was used to exclude oxygen from solutions containing partially reduced enzymes that would be oxidized by the oxygen. However, in the absence of oxygen, glucose oxidase can begin to reduce other electron acceptors, such as (6,7-Dihydrodipyrido[1,2-a:2',1'-c]pyrazinediium dibromide (6,7-DPD), which were used as a marker for when oxygen was thoroughly scrubbed from solution. Using steady-state kinetics and pseudo-first-order conditions, the cumulative results show that the initial rate of reaction increased linearly as concentration of 6,7-DPD increased, showing that an increase in substrate led to a first-order increase in the rate of substrate disappearance. It can be concluded from kinetic data that the rate of 6,7-DPD reduction was a significantly slower process than the rate of oxygen reduction to hydrogen peroxide. An induction period of ~430 seconds, that persisted until the concentration of oxygen diminished significantly, was shown to be independent of the concentration of 6,7- DPD. Over extended time periods glucose oxidase catalyzed reduction of 6,7-DPD by glucose proceeded via two distinct exponential phases, showing that the rate-determining step changed over time. One proposed explanation is that as the product lactone accumulates it inhibits Gox due to tight binding at the active site (product inhibition).

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Apr 24th, 10:30 AM Apr 24th, 11:45 AM

Glucose Oxidase as an Oxygen Scrubber

Union Wisconsin Room

Cytochrome C Nitrate Reductase is an anerobic enzyme that shows promise for study to generate raw material for fertilizers. Since the enzyme requires oxygen-free envrionments, hyper-anerobic environments were sought in an anerobic glove box using glucose oxidase to remove trace oxygen. Glucose oxidase catalyses the oxidation of glucose by oxygen to yield gluconolactone and hydrogen peroxide via a ping-pong bi-bi mechanism. The glucose/glucose oxidase pair was used to exclude oxygen from solutions containing partially reduced enzymes that would be oxidized by the oxygen. However, in the absence of oxygen, glucose oxidase can begin to reduce other electron acceptors, such as (6,7-Dihydrodipyrido[1,2-a:2',1'-c]pyrazinediium dibromide (6,7-DPD), which were used as a marker for when oxygen was thoroughly scrubbed from solution. Using steady-state kinetics and pseudo-first-order conditions, the cumulative results show that the initial rate of reaction increased linearly as concentration of 6,7-DPD increased, showing that an increase in substrate led to a first-order increase in the rate of substrate disappearance. It can be concluded from kinetic data that the rate of 6,7-DPD reduction was a significantly slower process than the rate of oxygen reduction to hydrogen peroxide. An induction period of ~430 seconds, that persisted until the concentration of oxygen diminished significantly, was shown to be independent of the concentration of 6,7- DPD. Over extended time periods glucose oxidase catalyzed reduction of 6,7-DPD by glucose proceeded via two distinct exponential phases, showing that the rate-determining step changed over time. One proposed explanation is that as the product lactone accumulates it inhibits Gox due to tight binding at the active site (product inhibition).