OXIDO-REDUCTASES May 11, 2018

Physical Biology

Assignment on:
 Oxido-reductases

Submitted to:
Dr.Habib Ullah Nadeem
(Dept.Of Bioinformatics &
Biotechnology)
Submitted By:

Muhammad Muneeb Nasir

Roll No.16642

BIT 2nd Semester Fall 2018

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 OXIDO-REDUCTASES May 11, 2018

Oxido-Reductases
An oxidoreductase is an enzyme that catalyzes the transfer of
electrons from one molecule, the reductant, also called the
electron donor, to another, the oxidant, also called the electron
acceptor. This group of enzymes usually utilizes NADP or NAD+
as cofactors.

Explanation:-
Oxidoreductases are a class of enzymes that catalyze oxidoreduction
reactions . Oxidoreductases catalyze the transfer of electrons from
one molecule (the oxidant) to another molecule (the reductant).
Oxidoreductases catalyze reactions similar to the following, A– + B
→ A + B– where A is the oxidant and B is the reductant .
Oxidorecuctases can be oxidases or dehydrogenases. Oxidases are
enzymes involved when molecular oxygen acts as an acceptor of
hydrogen or electrons. Whereas, dehydrogenases are enzymes that
oxidize a substrate by transferring hydrogen to an acceptor that is
either NAD+/NADP+ or a flavin enzyme. Other oxidoreductases
include peroxidases, hydroxylases, oxygenases, and reductases.
Peroxidases are localized in peroxisomes, and catalyzes the reduction
of hydrogen peroxide. Hydroxylases add hydroxyl groups to its
substrates. Oxygenases incorporate oxygen from molecular oxygen
into organic substrates. Reductases catalyze reductions, in most cases
reductases can act like an oxidases .

Oxidoreductase enzymes play an important role in both aerobic and

anaerobic metabolism. They can be found in glycolysis, TCA cycle,
oxidative phosphorylation, and in amino acid metabolism. In

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glycolysis, the enzyme glyceraldehydes-3-phosphate dehydrogenase

catalyzes the reduction of NAD+ to NADH. In order to maintain the
re-dox state of the cell, this NADH must be re-oxidized to NAD+,
which occurs in the oxidative phosphorylation pathway. Additional
NADH molecules are generated in the TCA cycle. The product of
glycolysis, pyruvate enters the TCA cycle in the form of acetyl-CoA.
During anaerobic glycolysis, the oxidation of NADH occurs through
the reduction of pyruvate to lactate. The lactate is then oxidized to
pyruvate in muscle and liver cells, and the pyruvate is further
oxidized in the TCA cycle. All twenty of the amino acids, except
leucine and lysine, can be degraded to TCA cycle intermediates. This
allows the carbon skeletons of the amino acids to be converted into
oxaloacetate and subsequently into pyruvate. The gluconeogenic
pathway can then utilize the pyruvate formed .

Reactions:-
For example, an enzyme that catalyzed this reaction would be an
oxidoreductase:
A– + B → A + B–
In this example, A is the reductant (electron donor) and B is the
oxidant (electron acceptor).
In biochemical reactions, the redox reactions are sometimes more
difficult to see, such as this reaction from glycolysis:
Pi + glyceraldehyde-3-phosphate + NAD+ → NADH + H+ + 1,3-
bisphosphoglycerate
In this reaction, NAD+ is the oxidant (electron acceptor), and
glyceraldehyde-3-phosphate is the reductant (electron donor).

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Classification:-
Proper names of oxidoreductases are formed as "donor:acceptor
oxidoreductase"; however, other names are much more common. The
common name is "donor dehydrogenase" when possible, such as
glyceraldehyde-3-phosphate dehydrogenase for the second reaction
above. Common names are also sometimes formed as "acceptor
reductase", such as NAD+ reductase. "Donor oxidase" is a special
case where O2 is the acceptor.

Quinones and Quinone Enzymes:-

Quinone oxidoreductases of the plasma membrane relate functionally
to the operation of a cell surface redox chain, where cytosolic
NAD(P)H is oxidized. Plasma membrane quinones serve as lipid-
soluble transmembrane shuttles to transfer the 2H+ + 2e- from
NAD(P)H to 1/2 O2 to form water. The reduction of 1/2 O2 is at the
expense of hydroquinone catalyzed by cell surface hydroquinone
oxidases. The first demonstration of a redox-related plasma
membrane enzyme was that of an NADH-ferricyanide reductase
observed with purified fractions of plasma membranes isolated from
rat liver. A plasma membrane location was subsequently confirmed
by electron microscope cytochemistry. Involvement in plasma
membrane electron transport was inferred from observations in which
ferricyanide and other impermeant oxidants were reduced by intact
cells. Quinone oxidoreductases, of the plasma membrane, have
physiological significance as components in support of plasma
membrane electron transport. Among these are the ECTO-NOX
proteins that comprise a family of NAD(P)H oxidases of plants and
animals that exhibit both oxidative and protein disulfide isomerase-
like activities that alternate and serve as terminal oxidases for plasma
membrane electron transport. A unique feature of the ECTO-NOX
proteins is that the two enzymatic activities they catalyze,

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hydroquinone (NADH) oxidation and disulfide–thiol interchange,

alternate within a 24-min period.

Biotransformation Reactions and their Enzymes

Bernard Testa1, Bernd Clement2, in The Practice of Medicinal
Chemistry (Fourth Edition), 2015

Other Oxidoreductases
Other oxidoreductases that can play a role in drug oxidation are
1. Monoamine oxidases, which are essentially mitochondrial
enzymes;
2. The broad group of copper-containing amine oxidases, which
contain diamine oxidase (DAO) and semicarbazide-sensitive
amine oxidases (SSAO);
3. The cytosolic molybdenum hydroxylases, namely aldehyde
oxidase and xanthine oxidoreductase, which exist in a
dehydrogenase form (XDH) and an oxidase form (XO);
4. Various peroxidases, such as eosinophil peroxidase (EPO),
myeloperoxidase (MPO) and thyroid peroxidase (TPO) (note
that several cytochrome P450 enzymes have been shown to have
peroxidase activity);
5. Prostaglandin G/H synthase, which is able to use a number of
xenobiotics as cofactors in a cooxidation reaction.
6. Dehydrogenases/reductases involved in reactions of oxidation
(dehydrogenation) and/or reduction (hydrogenation) are:
7. Alcohol dehydrogenases (ADH), which are zinc enzymes found
in the cytosol of the mammalian liver and in various
extrahepatic tissues;
8. Aldehyde dehydrogenases (ALDH), a large superfamily of
enzymes produced by nineteen human genes in eleven families
and thirteen subfamilies;

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9. The aldo-keto reductases (AKR), a complex superfamily of

enzymes, which includes aldehyde reductases (ALR) and
dihydrodiol dehydrogenase (DD);
10. Carbonyl reductases (CR) and quinone reductases (NQO);
11. Mitochondrial amidoxime reducing component (mARC).

Biocatalysis for Chiral Synthesis

Hyun-Dong Shin, ... Rachel R. Chen, in Bioprocessing for
Value-Added Products from Renewable Resources, 2007.

CHIRAL MOLECULES FROM ENZYMES

REQUIRING COFACTORS
Isolated enzymes
Using oxidoreductases as isolated enzymes necessitate in–situ
cofactor regeneration. The regeneration of pyridine nucleotides,
NAD(H) and NADP(H), is generally carried out by enzymatic
methods using a dehydrogenase or a NADH oxidase [42, 43].
Although much progress has been made, cofactor regenerations
remain an active area of research. New developments include
finding enzymes that are resistant to organic solvent, as most
substrates are not readily soluble in aqueous solution, and
organic-aqueous biphasic systems are more commonly used.

Novel enzymes are being sought for effectively regenerating

both reduced and oxidized cofactors. A recent and notable
development is a newly-discovered soluble pyridine nucleotide
transhydrolase (STH) from Pseudomonas fluorescens useful for
the regeneration of NAD and NADP. It was successfully used
for hydromorphone synthesis in both cell-free and whole-cell
systems [44]. STH can transfer reducing equivalents between
NAD(H) and NADP; thus an efficient cofactor recycling in the
presence of catalytic amounts of cofactors occurs in a cell-free
system, resulting in 84% high yield of hydromorphone. Another

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promising new enzyme, phosphite dehydrogenase, was

discovered from Pseudomonas stutzeri WM88 and can be used
for NAD regeneration [45]. This enzyme catalyzes the NAD-
dependent oxidation of phosphite to phosphate, and is useful for
the production of isotopically labeled products with a high
isotopic purity. Furthermore, the cofactor specificity of the
enzyme was successfully modified by using site-directed
mutagenesis; mutants were generated for the regeneration of
NADP. Due to the intrinsically large thermodynamic driving
force, the innocuous nature of phosphite and phosphate to
enzymes, and the low cost of phosphite, this phosphate–
phosphite dehydrogenase system may prove complementary to
the most widely used formate–formate dehydrogenase system.

Impressive progress has been achieved in recent years in

enzyme-coupled regeneration of oxidized cofactors. Particularly,
water-forming NADH oxidases were discovered from
Lactobacillus sanfranciscensis and Borrelia burgdorferi.

2NADH + O2 + 2H+ → 2NAD+ + 2H2O

The enzymes were cloned and heterologously expressed in E.
coli [46]. The enzyme from Lactobacillus sanfranciscensis
accepts NADPH with about 30% of the activity detected with
NADH, and hence may be used for regenerating both cofactors.
The enzyme was applied in the resolution of rac-1-
phenylethanol with ADH from L. brevis yielding
enantiomerically pure S-isomer [47]. Unlike regeneration using
lactate dehydrogenase or glutamate dehydrogenase, this method
does not generate byproducts other than water, but the oxygen
sensitivity of the protein remains a disadvantage.

Numerous literatures can be found documenting the

combination of an oxido-reductase and an enzyme for cofactor

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regeneration in chiral synthesis. Some reached preparative scale,

as shown in the following example. A biphasic system
containing 20% heptan and 80% aqueous buffer (pH 7.0) was
used for the asymmetric bioreduction of p-chloroacetophenone
with (S)alcohol dehydrogenase from Rodococcus erythropolis
under in situ-cofactor-recycling with a formate dehydrogenase
from Candida boidinii [48]. Interestingly, with this biphasic
system, reductions of poorly water-soluble ketones could be
carried out at higher substrate concentrations of 10–200 mM.
The resulting (S)-alcohols were formed with moderate to good
conversion rates, and with up to 99% ee.

Enantioselective reduction of C-4-substituted 3,5-

dixocarboxylates was reported using alcohol dehydrogenase
from Lactobacillus brevis (LBADH) over-expressed in E. coli
[49]. LBADH can recycle its cofactor by oxidation of 2-
propanol, as shown in Fig. 8. When 2-propanol was used as a
co-substrate, this enzyme reduced tert-butyl 3,5-dioxohexanoate
and tert-butyl 3,5-dioxoheptanoate on a preparative scale to the
corresponding Δ-hydroxy-β-keto esters, tert-butyl (R)-5-
hydroxy-3-oxohexanoate and tert-butyl (R)-5-hydroxy-3-
oxoheptanoate with 99.4% ee and 98.1% ee, respectively. These
esters are useful intermediates in the synthesis of
pharmaceuticals such as HMG-CoA reductase inhibitor, mevinic
acid.

Subclasses

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Oxidases transfer two electrons from the donor to oxygen,

forming hydrogen peroxide.

Cytochrome oxidase produces water molecules rather than

hydrogen peroxide by reducing the oxygen. Cytochrome P-450
catalyzes oxidation reactions in the liver and adrenal cortex,
helping to detoxify some substances by adding hydroxyl groups
that make the compounds more water-soluble and more
susceptible to further reactions. Unfortunately, at times this
process has the reverse effect because some relatively safe
molecules are converted into potent carcinogens. A group of
cytochromes serve as oxidation-reduction agents, converting the
energy of the oxidation process into the synthesis of adenosine
triphosphate (ATP), which makes the energy more available to
other reactions.
Oxygenases oxidize a substrate by incorporating oxygen to it.
Monooxygenases incorporate a single oxygen atom, and the
other oxygen is reduced by electrons from substrate to water.
Dioxygenases transfer two oxygen atoms in a molecule of O2 to
the substrate.
Peroxidases and Catalases etoxify hydrogen peroxide, which
acts as both electron donor and acceptor. Peroxidases and
catalases are Fe (III)-heme compounds that decompose
hydrogen peroxide and organic peroxides. The reactions seem to
proceed through Fe(IV) compounds with another unpaired
electron electron on the porphyrin, which becomes a radical
cation. Similar intermediates are also known in simpler
porphyrin molecules.

THE END