Bioenergetics Question B-14


 

 

This compound dissipates the electrochemical gradient between the intermembrane space and the mitochondrial matrix, consequently the electron transportation takes place but not the synthesis of ATP

 

a)     Antimycin B

b)     Carbon Monoxide

c)      Oligomycin

d)     Rotenone

e)     UCP

Inhibitors of the Electron Transport Chain


 

Answer to Question B-09

 

As described in a former post, the inhibitors of the Electron Transport Chain are substances that bind to some of the components of the ETC blocking its ability to change in a reversible form from an oxidized state to a reduced state.

 

This inhibition results in the accumulation of reduced forms before the inhibitor point, and oxidized forms of the components of the ETC downstream (ahead) the inhibition point.

 

 

Since energy is not released, the synthesis of ATP also stops. The most important known inhibitors of the ETC are Amytal, Rotenone, Antimycin A, CO, Sodium Azide, and Cyanides.

 

Amytal, a barbiturate, and Rotenone, a plant product used as insecticide and pesticide, block the ETC between NADH dehydrogenase (Complex I) and CoQ.

 

Consequently, they prevent the utilization of NADH as a substrate. On the contrary, electron flow resulting from the oxidation of Complex II is not affected, because these electrons enter through QH2, beyond the block.

 

The effect of Amytal has been observed in vitro, since the intoxication with amytal and other barbiturates in vivo affect mainly the CNS by acting on GABA-sensitive ion channels, an effect not related to the action of Amytal on Complex I.

 

Rotenone intoxications are very rare. In facts, some human tribes used to catch fishes by spreading plant extracts containing rotenone in the water, and this substance was easily absorbed by the fishes through the gills. These fishes were eaten later without notable side effects in humans, since rotenone is absorbed very difficult by the gastrointestinal tract. Usually, when taken in a concentrated form, irritating action in mucoses causes vomits.

 

It is interesting to note that Rotenone and MPTP (a neurotoxin), when administered in vein,  cause at the same time interference with the functioning of Complex I and a Parkinson-like disease. These substances affect primary neurons in substancia nigra; apparently the sequence is: impairment of Complex I, impairment of mitochondria metabolism, accumulation of free radicals, cell death, release of toxic compounds and destruction of other cells.

 

Antimycin A is an antibiotic produced by Streptomyces griseous that has been used as a piscicide for the control of some fish species. Antymicine A interferes with electron flow from cytochrome bH in Complex III (Q-cytochrome c oxidoreductase).  In the presence of this substance, cytochrome bH can be reduced but not oxidized, consequently,  in the presence of antimycin A cytochrome c remains oxidized, as do the cytochromes a and a3 that are ahead.

 

Carbon monoxide (CO) is responsible for more than 50 % of death by poisoning worldwide. It is colorless and odorless; high levels can result from incomplete combustion of fuels: engine and furnace exhausts are important sources. Tobacco smoking increases CarboxyHb levels.

 

Carbon monoxide intoxication causes impaired oxygen delivery and utilization at the cellular level. The affinity of Hb for CO is almost 300 times higher than for Oxygen. An environment in which there is 100 ppm of CO is enough to form 16 % carboxyhemoglobin. The situation is worsen since the binding of CO to one of the Hem groups of Hemoglobin increases the affinity of the other three Hem groups for Oxygen, so the delivery of Oxygen to  tissues is very affected. The brain and the heart, that has a high Oxygen consumption, are the most affected. Myoglobin has even a greater affinity for CO than Hemoglobin. As a consequence of the binding of CO to these molecules, the heart functioning is very impaired and the patient presents sever hypotension. As described above, this intoxication is an important cause of death worldwide.

 

The affinity of respiratory chain components for CO is lower than for Oxygen,

but since the clinical status does not correlate very well with the carboxyhemoglobin levels, it is considered that the inhibition of Cytochrome Oxidase by CO also plays a role in CO intoxication. CO binds to the reduced form of iron in Hem groups (Fe++) in cytochrome Oxidase

 

On the contrary, in cyanide intoxication the inhibition of the respiratory chain has a primary role. Intoxication by cyanide can be seen relatively frequent in patients with smoke inhalation from residential or industrial fires. Also in persons related professionally with cyanide or derivatives in certain industries.  Intentional poisoning can be seen in suicidal persons with access to cyanide compounds. Cyanide affects practically all metalloenzymes, but its principal toxicity derives from the binding to the Fe+++ in the Hem groups in cytochrome Oxidase, inhibiting the functioning of the Electron Transport Chain. As a consequence, redox reactions in the respiratory chain will stop, energy will not be released, proton pumps will not function, so they will not return through Complex V, and the production of ATP will cease (Related question here).

 

Azides have an action on the respiratory chain very similar to cyanide, inhibiting the Hem groups of cytochromes in Cytochrome Oxidase (Complex IV). Azides are used as propellants in airbags, in detonant (explosive) industry and as preservative of sera an reagents. Some cases of azide intoxication in humans have been reported.

 

You can find more information about these inhibitors of the Electron Transport Chain in these links:

 

Antimycin A: toxicity, ecological toxicity and regulatory information

 

 

Risk assessment for Piscicidal Formulations of Antimycin

 

Leybell, I: Toxicity: Cyanide

 

Cyanide poisoning

 

 

Azide Toxicity

 

Sodium Azide Toxicity effects

 

Shochat, G.N.: Toxicity, Carbon Monoxide

 

 

Substances which interfere with the Respiratory Chain: Overview


Recalling:

 

Respiratory Chain:

 

Set of reaction that transfers the Hydrogen and electrons from reduced cofactors to Oxygen, obtaining water and using the released energy for the synthesis of ATP

 

 

Sometimes, the terms Electron Transport Chain and Oxidative Phosphorylation are used indistinctly to refer to the Respiratory Chain. We prefer to use Respiratory chain for the whole process, in which we can distinguish two different parts:

 

         Electron transport chain: Set of reaction through which the Hydrogen and electrons are transfer from reduced cofactors to Oxygen, obtaining water and releasing energy. It allows that the energy derived from redox reactions be released little by little with a better use and without damaging the cell. It involves the transfer of electrons from complex I (or II) to Oxygen. (An excellent animation here)

          Oxidative phosphorylation: It’s the biosynthesis of ATP from ADP + (P) using the energy released in the electron transport chain. It occurs in Complex V (ATP synthase). The coupling of ATP synthesis and the energy released in the ETC is indirect (Animation here)

 

Substances that interfere with the Respiratory Chain function can be classified as:

 

Inhibitors of Electron Transport Chain:

Substances that bind to some of the components of the ETC blocking its ability to change in a reversible form from an oxidized state to a reduced state. It results in the accumulation of reduced forms before the inhibitor point, and oxidized forms of the components of the ETC ahead the inhibition point. Sites of action of some inhibitors (and some artificial electron acceptors) can be found in this graphic

Since energy is not released, the synthesis of ATP also stops.

Ex: Amital, Rotenone, Antimycin A, CO, Sodium Azide,  CN

 

Inhibitors of Oxidative Phosphorylation:

These compounds bind to the Complex V (ATP synthase), impeding the synthesis of ATP, by inhibiting the return of protons to the matrix. Since this process is coupled to the ETC these inhibitors also stop the function of the ETC.

         Ex Oligomycin

 

Uncouplers of respiratory chain:

Substances that dissipate the electro-chemical gradient by facilitating the entrance of protons to the matrix using “shortcuts” and by that reason the electron transport takes places but not the oxidative phosphorylation.

         Ex. 2,4 dinitrophenol

         Thermogenin – UCP-1, DNP

 

Inhibitors of ATP/ADP exchange

ATP is produced in the mitochondria but is used all over the cell, so it should be allowed to go from the production site (mainly, mitochondria) to the locations where it is going to be used. At the same time, the ADP resulting from the use of ATP all over the cell should be allowed to enter the matrix in order to be used in the formation of more ATP.  The inhibitors of ATP/ADP exchange inhibit the translocase that allows the entrance of ADP to the matrix and the output of recently synthesized ATP outside of the mitochondria.

         Ex Atractyloside

 

More information about these substances that interfere with the Respiratory Chain will be included in future posts dedicated to each specific group.

 

 

Question about Bioenergetics (B-09)


 

 

Intravenous nitroprusside rapidly lowers elevated blood pressure through its direct vasodilating action. It is use to treat severe hypertension in acute care situations.  It must be administered cautiously, since it is converted to cyanide and its use sometimes is checked for toxicity. Certainly this is advisable, since cyanide act in the Respiratory chain as:

 

a)     Inhibitor of the ATP/ADP exchange

 

b)     Inhibitor of the ATP synthase

 

c)      Inhibitor of Cytochrome Oxidase

 

d)     Inhibitor of Complex III

 

e)     Inhibitor of the NADH.H dehydrogenase

 

f)       Inhibitor of translocases

 

g)     Ionophor agent increasing the permeability of the inner membrane

 

h)    Uncoupler of oxidative phosphorylation from electron transfer chain

 

 

 

Answer

 

 

 

A: More questions about Bioenergetics (B-03 to B-05)


Original Questions

Answer to Bioenergetics Question B-03

                                                                       

Answer: (d)

 

Answering this kind of questions is very easy. It just requires recalling the composition and organization of the Respiratory Chain:

 

 

 

NADH.H+ is accumulated. Observe that reductions equivalents (Hydrogens) from NADH.H+ enter the electron transport chain through Complex I (NADH dehydrogenase). It means that Complex I is not working properly.

Hydrogens from FAD can not enter the electron transport chain either. It means that Complex II (succinate dehydrogenase) and other enzymes related to the Electron Transport Chain hat use FAD as cofactor can not be dehydrogenated.

 

Coenzyme Q (Ubiquinone) is also accumulated in a reduced form.

 

Which compound is in an oxidized state? Cytochrome c.

 

It is obvious then that there is a problem between CoQ, that remains reduced, and Cytochrome c, that remains oxidized.  (Of course, since Coenzyme Q remains reduced, it can not accept more electrons from FADH2 or from NADH.H+ and these cofactors remain in the reduced state.)

 

Observing the organization of the electron transport chain, we can say that this new inhibitor probably is affecting Complex III.

 

Antimycin A is known to block the transfer of electrons between two components of Complex III, (Cytocrom b and c1), so its effect would be very similar to the effects described in the question for the new inhibitor.

 

 

Answer to Bioenergetics Question B-04:

 

Answer (e)

 

Once again, observe the organization of the respiratory chain.

 

X represents Complex III

 

 

Answer to Bioenergetics Question B-05:

 

Answer: (e)

 

ATP synthase is called Complex V of the Respiratory chain (observe the position in the graphic).

During the spontaneous flow of electrons through the ETC (from the component with the most negative to the component with the most positive redox potential), energy is released. This energy is used to pump protons from the mitochondrial matrix to the intermembrane space.

 

This accumulation of protons in the intermembrane space creates an electrical potential (positive in the outside of the internal membrane) and a pH gradient (outside the internal membrane is more acid, since there are more free H+).

 

These protons tend to return spontaneously to the matrix, driven by the electrical and the chemical gradient that have been created through their pumping across the membrane.

 

ATP synthase synthesize ATP from ADP and (P) using the energy released from the spontaneous return of protons from the intermembrane space to the matrix.

 

That is why it is said that the energetic coupling between the ETC and the Oxidative Phosphorylation is indirect.

 

Notes:

 

Sometimes the terms Electron Transport chain and Respiratory chain are use indistinctly. However, a more strictly use restricts the term Electron Transport Chain to the components of the Respiratory chain responsible of the transport of reduction equivalents from reduced cofactors to Oxygen, forming water, and releasing energy in the process.

 

Oxidative Phosphorylation is the process of biosynthesis of ATP using the energy that is released in the respiratory chain. (This energetic coupling is indirect, as described before)

 

The concept of Respiratory chain then would include the concepts of Electron Transport Chain and Oxidative Phosphorylation, and should be described as the set of reactions through which reduction equivalents are transferred from reduced cofactors to Oxygen releasing energy that is used for the biosynthesis of ATP.

 

The integration between the Electron Transport Chain and the Oxidative phosphorylation, forming the Respiratory chain, is clearly seen in this short movie

 

This animation also illustrates this relationship:

Voet, Voet, Pratt: Fundamentals of Biochemistry

The coupling of Electron Transport chain and ATP Synthesis

 

 

More Questions About Bioenergetics


 

 

Question B-03

 

A newly discovered inhibitor was found to cause the accumulation of NADH.H+, FMNH2, reduced CoQ, but not of reduced cytochrome c. It is most likely to be:

 

a)     an uncoupler of oxidative phosphorylation

b)     affecting a component of complex I

c)      affecting a component of complex II

d)     affecting a component of complex III

e)     affecting a component of complex IV

f)       affecting a component of complex V

 

 

Question B-04

 

 

In the electron transport scheme below,

Complex I-> CoQ  ->  X ->cit c  -> Complex IV -> O2 

X represents which of the following?

 

a)     NAD

b)     FMN

c)      Complex II

d)     ATP synthase

e)     Complex III

f)       Complex V

g)     FAD H2

 

 

Question B-05

 

The synthesis of ATP in the Respiratory Chain is performed by:

a)     Complex I

b)     Complex II

c)      Complex III

d)     Complex IV

e)     Complex V

  

Answers