Biological Oxidation

Oxidation is defined as the loss of electrons and reduction as the gain of electrons. This
may be illustrated by the interconversion of ferrous ion (Fe2+) to ferric ion (Fe3+).

The electron lost in the oxidation is accepted by an acceptor which is said to be reduced.
Thus the oxidation-reduction is a tightly coupled process.
The general principle of oxidation-reduction is applicable to biological systems also. The
oxidation of NADH to NAD+ coupled with the reduction of FMN to FMNH2 is illustrated

In the above illustration, there are two redox pairs NADH/NAD+ and FMN/FMNH2. The
redox pairs differ in their tendency to lose or gain electrons.
Biological Oxidation
The transfer of electrons from reduced coenzymes through the respiratory chain to oxygen
is known as biological oxidation. Energy released during this process is trapped as ATP. This
coupling of oxidation with phosphorylation is called oxidative phosphorylation.
Redox potential (E0)
The oxidation-reduction potential or, simply, redox potential, is a quantitative measure of
the tendency of a redox pair to lose or gain electrons. The redox pairs are assigned specific
standard redox potential (E0 volts) at pH 7.0 and 25°C.
The more negative redox potential represents a greater tendency (of reductant) to lose
electrons. On the other hand, a more positive redox potential indicates a greater tendency (of
oxidant) to accept electrons. The electrons flow from a redox pair with more negative E0 to another
redox pair with more positive E0.

ENZYMES & COENZYMES INVOLVED IN BIOLOGICAL OXIDATION

All the enzymes participating in biological oxidation belong to the class oxidoreductases.
These are further grouped into four categories
1. Oxidases
2. Dehydrogenases
3. Hydroperoxidases
4. Oxygenases.
1. Oxidases:
These enzymes catalyse the elimination of hydrogen from the substrates which is accepted
by oxygen to form mostly water,
E.g. cytochrome oxidase, tyrosinase, monoamine oxidase (H2O2 formed instead of H2O).

Cytochrome oxidase, the terminal component of electron transport chain, transfers

electrons (obtained from the oxidation of substrate molecules by dehydrogenases) to the final
acceptor, oxygen.
Some flavoproteins containing FAD or FMN also belong to the category of oxidases.
e.g., L-amino acid oxidase (FMN), xanthine oxidase (FAD).

2. Dehydrogenases:
As the name indicates, these enzymes cannot utilize oxygen as hydrogen acceptor. They
catalyse the reversible transfer of hydrogen from one substrate to another and, thus, bring about
oxidation reduction reactions.
 a) Aerobic dehydrogenases
They contain flavoproteins (FMN, FAD) as coenzyme. They transfer hydrogen
atom from the substrate to oxygen via flavin carriers and are called as aerobic
dehydrogenases. They are also capable of transferring hydrogen to acceptors other than
oxygen.
b) Anaerobic dehydrogenases
Perform 2 main functions.
 Transfer hydrogen from one substrate to another in a coupled oxidation reduction reaction.
 As a components of ETC, it use coenzyme- riboflavin and nicotinamide as hydrogen
carrier.

There are a large number of enzymes belonging to dehydrogenase group

 NAD+ dependent dehydrogenases
E.g. alcohol dehydrogenase, glycerol 3-phosphate dehydrogenase.
 NADP+ dependent dehydrogenases
E.g. HMG CoA reductase, enoyl reductase.
 FMN dependent dehydrogenases
E.g. NADH dehydrogenase.
 FAD dependent dehydrogenases
E.g. succinate dehydrogenase, acyl CoA dehydrogenase.
 The cytochromes :
All the cytochromes of electron transport chain (b, c1 and c) except the terminal
cytochrome oxidase (a+a3) belong to this group.

3. Hydroperoxidases:
Hydrogen peroxide is the substrate for these enzymes. There is a constant production of
H2O2 in the reactions catalysed by the aerobic dehydrogenases. The harmful effects of H2O2 are
prevented by hydroperoxidases.
E.g. peroxidase and catalase.
 Peroxidases utilizes H2O2 as oxygen donor but oxygen acceptor is a molecule other than
H2O2.
E.g.: Glutathione peroxidase
Catalases is a unique enzyme and utilizes H2O2 as both donor and acceptor of oxygen
(electrons).

4. Oxygenases:
This group of enzymes catalyses the direct incorporation of oxygen into the substrate
molecules.
 Dioxygenases (true oxygenases) :
They are responsible for the incorporation of both the atoms of oxygen (O2) into
the substrate
E.g. homogentisate oxidase, L-tryptophan pyrrolase.

 Mono oxygenases (mixed function oxidases)

They catalyse the incorporation of one atom of oxygen (1/2 O2) while the other oxygen
atom is reduced to H2O. NADPH usually provides the reducing equivalents.
E.g. cytochrome P450 mono oxygenase system of microsomes is responsible for the
metabolism of many drugs (amino pyrine, morphine, aniline etc.) and biosynthesis of steroid
hormones (from cholesterol). The action of Cyt P450 is depicted here.
 Electron Transport Chain
A series of compounds that transfer electrons from electron donors lo electron acceptors
through oxidation and reduction carried out simultaneously & couples this electron transfer with
the transfer of protons (H+ ions) across a membrane. The process generates a transmembrane
electrochemical gradient across the membrane & derives the synthesis adenosine tri phosphate
(ATP). The chain is also known as respiratory chain. In the electron transport chain electrons flow
from electro negative potential to electro positive potential.
Location:
The ETC is functioning inside the mitochondria, specifically in the inner mitochondrial
membrane where its serves as the site of oxidative phosphorylation in eukaryotes. In the inner
membrane, electrons from NADH & FADH2, pass through ETC to oxygen which is reduced to
water.
Organization of ETC
(i) In the ETC, the electrons are transferred from NADH to a chain of electron carriers.
The electrons flow from the more electronegative components to the more
electropositive components.
(ii) All the components of ETC are located in the inner membrane of mitochondria.
(iii) There are four distinct multiprotein complexes named as complex I, II, III & IV. These
are connected by two mobile carriers co-enzyme Q & cytochrome C.
Complex I
It’s known as NADH- CoQ reductase or NADH dehydrogenase complex. The complex
tightly bound to the inner membrane of mitochondrion. It contains a flavoproteins consisting FMN
as prosthetic group and an iron- sulphur protein (Fe-S). NADH is the proton donor& FMN accepts
and then get reduced to FMNH2. Two electrons and one hydrogen ions are transferred from NADH
to the flavin prosthetic group of the enzyme.
NADH + H+ + FMN → FMNH₂ + NAD+
The electrons from FMNH2 are transferred to Fe-S. The electrons are transferred to CoQ
(ubiquinone). Thus totally a pair of electrons are transferred to CoQ from NADH by this complex.
Flavoproteins
The enzyme NADH dehydrogenase (NADH- coenzyme Q reductase) is a flavoprotein with
FMN as the prosthetic group. The coenzyme FMN accepts two electrons & a proton to form
FMNH₂.
NADH dehydrogenase is a complex enzyme closely associated with non-heme iron
proteins or iron-sulfur proteins. In this, 4 protons are pumped out from mitochondria.

COMPLEX II: Succinate-CoQ-Reductase

• The electrons from FADH2 enter ETC at the level of Co Q.

• Succinate DH is an enzyme found in inner mitochondrial membrane.
• It is also a flavoprotein with FAD as coenzyme.
• The 3 major enzyme systems that transfer their electrons directly to ubiquinone are:

a. Succinate dehydrogenase
b. Fatty acyl CoA dehydrogenase
c. Mitochondrial glycerol phosphate dehydrogenase.

Iron-sulfur Centers (clusters)

Iron-sulfur centers (Fe-S) are prosthetic groups containing 1-4 iron atoms. Iron-sulfur (Fe-
S) proteins exist in the oxidized (Fe3+) or reduced (Fe2+) state. Iron-sulfur centers transfer only one
electron, even if they contain two or more iron atoms. Fe-S participates in the transfer of electrons
from FMN to coenzyme Q.
Coenzyme Q
It is also known as ubiquinone. It is a quinone derivative with isoprenoid side chain.
Mammalian tissues possess a quinone with 10 isoprenoid units which is known as coenzyme Q10.
The ubiquinone is reduced successively to semiquinone & finally to quinol.
It accepts a pair of electrons from NADH or FADH2 through complex I or complex II
respectively. 2 molecules of cytochrome c are reduced. The Q cycle facilitates the switching from
the 2 electron carrier ubiquinol to the single electron carrier cytochrome c. This is a mobile carrier.
COMPLEX III: Cytochrome Reductase
This is a cluster of iron-sulphur proteins, cytochrome b & cytochrome c1, both contain
heme prosthetic group. Cytochromes are conjugated proteins. Consists of a porphyrin ring with
iron atom.
The iron of heme in cytochromes is alternately oxidized (Fe³+) & reduced (Fe²+) Which is
essential for transport of electrons in the ETC. In this, 4 protons are pumped out. The electrons
transported from coenzyme Q to cytochromes b, c1, c, a & aз.
The property of reversible oxidation- reduction of heme iron present in cytochromes allows
them to function as effective carriers of electrons in ETC.
Cytochrome C
It is a small protein containing 104 amino acids & a heme group. It is a loosely bound to
inner mitochondrial membrane & can be easily extracted.
COMPLEX IV: Cytochrome Oxidase (Cytochrome a & aз)

Contains cytochrome a and cytochrome a3, which is the terminal component of ETC it is
tightly bound to inner mitochondrial membrane.
Cytochrome oxidase is the only electron carrier, heme iron of which can directly react with
molecular oxygen. It also contains copper that undergoes oxidation-reduction during transport of
electrons and 2 protons are pumped out.
In the final stage of ETC, the transported electrons, the free protons & the molecular
oxygen combine to produce water.

Sources of Electrons

Electrons donors: NADH & FADH2

NADH: It is produced in the following reactions:

• Pyruvate dehydrogenase complex: It transfers electrons from pyruvate to NAD+

• α-ketoglutarate DH: It transfers electrons from alpha-ketoglutarate to NAD+
• Isocitrate DH: It transfers electrons from isocitrate to NAD+
• Malate DH: It transfers electrons from malate to NAD+
• Hydroxy acyl CoA DH: It transfers electrons from hydroxy acyl CoA to NAD+
FADH2: FAD is tightly bound to enzymes called flavoproteins. It is produced in the following
reactions:

• Succinate DH (complex II): It transfers electrons from succinate to FAD.

• Glycerol 3-Phosphate DH: It transfers electrons from glycerol 3-P to FAD.
• Fatty acyl CoA DH: It transfers electrons from fatty acids to FAD.

• FADH2 donates electrons to coenzyme Q.

 Inhibitors of electron transport chain
Many site-specific inhibitors of ETC have contributed to the present knowledge of
mitochondrial respiration. Selected examples of these inhibitors are given below:

The inhibitors bind to one of the components of ETC and block the transport of electrons.
This causes the accumulation of reduced components before the inhibitor blockade step and
oxidized components after that step.
The synthesis of ATP (phosphorylation) is dependent on electron transport. Hence, all the
site-specific inhibitors of ETC also inhibit ATP formation. Three possible sites of action for the
inhibitors of ETC are identified
1. NADH and coenzyme Q: Fish poison rotenone, barbituate drug amytal and antibiotic
piercidin A inhibit this site.
2. Between cytochrome b & c1: Antimycin A — an antibiotic, British antilewisite (BAL) — an
antidote used against war-gas—are the two important inhibitors of the site between cytochrome b
and c1.
3. Inhibitors of cytochrome oxidase: Carbon monoxide, cyanide, hydrogen sulphide and azide
effectively inhibit cytochrome oxidase. Carbon monoxide reacts with reduced form of the
cytochrome while cyanide and azide react with oxidized form.
 Cyanide poisoning: Cyanide is probably the most potent inhibitor of ETC. It binds to Fe3+ of
cytochrome oxidase blocking mitochondrial respiration leading to cell death. Cyanide
poisoning causes death due to tissue asphyxia (mostly of central nervous system).
 Uncouplers