https://www.nature.com/articles/nrn1767

http://diabetes.diabetesjournals.org/content/51/9/2749.short

Hi Dewan,

As a consequence of their regulated proton channel/transport mechanism, UCPs are capable of dissipating the proton (H+) gradient generated by the mitochondrial respiratory chain. In other words, when the UCPs are active, they allow the protons to return to the matrix side, diminishing the proton-motive force and thus decreasing the ATP synthesis yield. In this case, the proton gradient is not used to do biochemical work (ATP synthesis, protein import, metabolite transport, etc.). Instead, energy is lost as heat, which from the thermodynamic point of view is "useless" energy. The respiratory chain will pump protons continuously, transferring electrons to oxygen, but most of the translocated protons will come back immediately through the UCP in a futile cycle.

However, UCPs activity is useful in some species for thermogenesis. For example, when some plants are exposed to cold temperatures, they express or activate UCPs decreasing the ATP synthesis but using the energy lost as heat to warm up. Other classical example is the UCP1 (thermogenin), which was discovered in the brown adipose tissue.

Dear Alfredo,

I wonder how the movement of H+ back to the matrix is able to generate heat in physical sense.

Interesting issue... what if by dissipating proton gradients, in other words, increasing entropy, the system converts this free energy into enthalpy. It sounds weird because it looks like adiabatic compression (gas), but here we don't have any gas particle.

Dear AnthonyKi, dear Omar

I'm happy to follow up on this question and I'm sure we can get a better understanding of this "uncoupling" process together. Let me elaborate more about this process.

First of all, I'd like to point out that the isolated ability of the UCPs letting the protons (H+) flowing back to the mitochondrial matrix does not explain by itself the energy loss as heat. We have to integrate its role alongside the oxidative phosphorylation (OXPHOS) pathway and other related metabolic processes; such as the TCA cycle.

In this regard, just to quick remind you guys that OXPHOS involves the respiratory chain complexes and the ATP synthase working together in a "coupled" fashion. What connects these two processes is the electrochemical gradient of H+; i.e. proton-motive force. This proton-motive force is canonically described as the ATP synthesis driving force; however, it is often forgotten that the energy accumulated as H+ gradient is not fully interconverted into chemical energy (e.g. ATP synthesis, metabolite transport, protein import, etc.); i.e. a big part of this energy is simply lost as H+ leaking.

I know that the inner membrane is thought to be impermeable to H+ and in a high degree it does; but, in real life, it is not 100% impermeable, meaning that the H+ may cross the membrane back to the matrix, following the thermodynamic driving forces; i.e. matrix side is negatively charged. Within an intrinsic leak in the system, how is the proton concentration maintained high enough to build proton-motive force? The answer is a sustained activity of the respiratory chain pumps. In order to generate the H+ gradient, the respiratory chain has to transfer electrons to the Oxygen continuously reaching a “steady-state”. The latter does depend on the usage of the proton-motive force. For example, if the ATP synthesis is high, the proton-motive force decreases thus increasing the activity of the respiratory chain to restore and keep up the proton gradient. In this scenario, a considerable amount of H+ are used by the ATP synthesis to drive phosphorylation, but at the same time we have the intrinsic H+ leak, which explains again why the efficiency of the system is not 100%; i.e. ca. 30% of the total free energy from the NADH oxidation is actually used for the synthesis of ATP. It is very useful to imagine the OXPHOS system as an electrical circuit, the respiratory chain is the battery that drives electrical charges flux (in this case H+ translocation) and the ATP synthase uses this energy to produce ATP (as a lamp uses electricity to get light); the intrinsic proton leaks would be seen as the electrical energy loss as heat (as it also occurs in the lamp). Again, heat is a form of energy which is “disordered” and useless to do effective work; i.e. it cannot be converted completely into work, regardless the type (1st Law of Thermodynamics).

Now, please imagine the scenario where the UCP is active and turns into a protein-mediated H+ leak, besides the intrinsic leak. The latter results in complete dissipation of the proton-motive force simply because this system offers less resistance (as in an electrical shortcut). Then, what happens here? The gradient is now gone, but the respiratory chain and the ATP synthase are still there and working. On the one hand, the respiratory chain is not restricted anymore by the “steady-state” of the proton gradient; therefore, it now works at full speed because there are still electron donors (NADH, succinate, etc.) coming from catabolic pathways, Oxygen and the transmembrane potential is very low; i.e. the pumps are not restricted by the proton-motive force. The matrix protons are still translocated by the respiratory pumps; however, these protons are not accumulated at the intermembrane space since the leaks prevent this and rapidly equilibrate the protons in both compartments. Thus, the “battery” is working, moving the protons, but no gradient builds up, which results in an energy loss as heat; i.e. energy cannot disappear, it has to be transformed and, in this case, it is not converted into any kind of work (useful energy). In addition, the ATP synthase is not driven anymore by the proton-motive force and no ATP synthesis can get place. In this scenario, the enzyme changes the catalytical reaction direction and now turns into an ATPase capable to hydrolyze ATP to pump H+ aiming to generate proton-motive force, which again will not be possible due to the leaks. ATP hydrolysis is spontaneous and fast, the energy released during this step results in no useful energy conversion, thus generating heat.

Omar Porras in biochemical systems the concentration of the solutes are often at a very diluted concentration; therefore these particles behave as gas particles and the thermodynamics describing these processes are totally based on the classic and gas-related equations. However, the heat resulting from the UCP activity is mainly coming from the futile energy cycle of the uncoupled OXPHOS activity.

Let me know what you guys think and hope these lines are useful!

Best, Alfredo

Well, nice descritption Alfredo.

Still, I have the same fundamental question. If UCP appears in the mitochondrial picture, a loss in the electrochemical gradient will occur, and the situation will be operating at steady-state, which means that heat is not depending on proton gradient; instead, it depends on electron transfers through the mitochondrial complexes. Taking the electrical model, electrical resistance generates heat, but here UCP does exactly the opposite; it is a proton permeant hole, so, for me, it is not clear the analogy.

Going to the fundamental models to assign heat to molecular phenomena, all of them claim for intramolecular vibrational energy (fluorescence, electron relaxation, etc.). But here, in the mitochondria, you have a conserved strategy, which is the expression of UCP, to generate heat. This point aims to something changes in the system that now, in the presence of UCP, more heat is produced, likely at the cost of ATP production.

Well, I accept that is not de dissipation of protons that produces heat, but then, is that the whole electron transfer complexes vibrate much more when a proton leak is on the inner membrane? there are 2 celsius degrees between brown fat and white fat. (Cell metabolism 19, 302-309, 2014).

The big issue here is to determine if we are able to define how the energy (some people like to use chemical, bonds, etc.) is converted or re-directed to produce heat. HEAT is only understood as vibrational energy, whatever the scale of observation?

Cheers, OP.

Dear Omar!

Please find attached a figure from the book of Nicholls and Ferguson (Bioenergetics) showing the analogy of the OXPHOS system with an electrical circuit to better illustrate what I've tried to explained previously. Although the activity of UCPs is often represented as a simple H+ leak, it should be noted that UCPs belong to the superfamily of mitochondrial carriers which have a very conserved 6 transmembrane helices structure. Thus, UCPs are not simple “holes” or “pores”, not even ion channels, they’re transporters and their molecular mechanisms involve conformational changes to allow the movement of H+ (please check this review for extra details: New advance in adaptive thermogenesis: UCP1 and beyond; https://www.cell.com/action/showPdf?pii=S1550-4131%2818%2930679-X).

The membrane itself is an organelle with electrical properties; i.e. it has intrinsic resistance, capacitance and inductance. Values for the latter depend of course on the type of membrane and the ion/solutes involved. The intrinsic proton leak has a very low resistance when compared with the proton translocation via the OXPHOS complexes; still this resistance value is neither zero nor neglectable. Proteins mediating ion and/or solutes transport may also have these properties and a lower conductance (1/R) than the phospholipids bilayer.

Regarding the point you mentioned about heat, I have some comments:

A) The role of UCPs in thermogenesis result from its OXPHOS uncoupling mechanism. They dissipate proton-motive force decreasing the ATP yield and pretty much eliminating the limiting step of the respiratory chain, which is the transmembrane potential value. In other words, the higher the proton-motive force is, the lower the respiration is. I suggest the analogy of a car’s engine… the engine is the respiratory chain, the fuel is the electrons, the tires are the ATP synthase, and the pistons and mechanical clutch system might be the protons and the gradient. The transmission of energy only occurs when the engine is clutched mechanically to the tires (I’m trying to be minimalistic and probably the example is not perfect, I've tried my best). In OXPHOS, ATP production is "clutched electrochemically" to the respiratory chain through the proton gradient.

B) The respiratory chain always releases heat, no matter what. The energy interconversion from the redox reactions to proton translocation is not 100% efficient; so, the lost energy is converted and released as heat (involving vibrations, molecular movement, conformational changes, entropy, etc). Therefore, the uncoupling action of the UCPs only accelerates the respiratory activity, which generates more heat and faster than in the “coupled” scenario. UCPs declutch the respiratory chain (engine) from the ATP synthesis (tires), but doesn’t prevent the respiration flux (fuel is continuously burned and pistons (protons) are moving). What happens if you push the accelerator pedal without clutching? The engine burns more fuel and the energy is lost moving the pistons in a futile fashion and more heat is hence released.

C) In most tissues, UCPs are expressed and tightly regulated by fatty acids and purine nucleotides; the fact they’re expressed doesn’t mean they’re fully active. This means that UCPs activity may only decrease the yield of ATP but not preventing its synthesis completely. It should not be a black and white situation, UCPs are quite regulated and integrated into the complex mitochondrial physiology. And you can tell me if 2 degrees does not represent a considerable temperature change; we feel terrible with a fever of 38-39 °C, isn’t it?

Kind regards,

Alfredo

Thanks, Alfredo,

Actually, I think 2 degrees is a huge amount of energy. By the way, I quite close with the movement of ion by channels as marvelous molecular machines, I followed the work of two great electrophysiologists of Drs. Mackinnon and Ramon Latorre, masters of potassium channels structure-function, so when I used the term "holes" were only a simplification.

Still is pending the big question. You state the following sentence... the respiratory chain generates heat... how? like an electrical circuit? good.. electrical circuits generate heat thanks to electrical resistance, which is the opposite of conductance. Now, UCP increases the conductance of protons. I think that mechanical analogy makes me think in shear stress force, it doesn't help to understand.

still, how many ways to generate heat do exists in the universe?

Omar Porras, the respiratory chain catalyzes a series of redox reactions to transfer electrons from NADH to Oxygen. These reactions involve multiple protein complexes and redox centers. Although the majority of these partial electron transferences are spontaneous, they don't occur at the same rate or following the same routes. As an example, complex I catalyzes only the electron transfer from NADH to ubiquinone, transferring first two electrons to its FMN site and then one electron at a time through a series of FeS clusters until they reach to the ubiquinone. Thermodinamically speaking, this overall process goes downhill but not all the semi-steps occur like this, you have some intermediate steps which might have a higher energy barrier than others. In this regard, the slow steps and low redox potentials are the ones offering a higher resistance, electrons are moving, hence it works in a way like an electrical circuit. The energy that is released during all these electron transfer steps (regardless the complex and part of the respiratory chain) has to be interconverted in some other type of energy; in this case in both chemical/vectorial work and heat. In the first case, only the complexes that are proton pumps (I, III and IV) couple the energy released from the electron transfer to the translocation of protons across the membrane, in other words these complexes do useful biochemical work. In the second case, however, the partial steps which are not directly involved in driving proton translocation release the energy simply as heat. Why? because if the energy could not be completely transformed into work, it interconverts into heat (1st law again... U = W + Q).

The fraction of energy which is now accumulated as potential energy within the proton gradient will get another energy loss as heat when the protons flow back across the membrane or the UCP and not via the ATP synthase (or other biochemical process, such as metabolite antiport/simport). In the scenario where the UCP is active and the proton gradient is dissipated, the interconverted energy coming from the redox reactions and stored originally as the proton gradient is either low or totally lost, but again this energy can't simply disappear, so, it is then converted and released as heat.

About your last question, I think we can spend quite some time to ellaborate and probably a physicist can explain better this, but from the theory... all kinds of energy transformations are capable to generate heat. In this context, breakdown of metabolites by the mitochondrial pathways, on the one hand, release energy which is transiently stored in electron carriers and then interconverted into proton-motive force resulting in the synthesis of ATP and oxidation of carbon sources to CO2 and H20 plus the lost energy fraction as heat (entropy). We're literally burning nutrients to store now this energy in biochemical work and since it's impossible to convert and use all this energy, the rest is simply gone as heat.

My pleasure discussing this with you ;)

Alfredo

Well, it is fascinating the way the energy moves in a myriad of reactions. I know that all physic phenomena, including biological processes, nominate heat as a by-product, as a constant at the end of the equation. However, here the big issue is that the arrangement of redox reactions confined in a close space was selected to generate heat. Remember that there are many other biochemical and biophysics processes that occur in other membranes as well, however, it is on mitochondrion where the heat seems to be generated with high efficiency, despite to be a "by-product"...

We should write a commentary about this fantastic topic.... what do you think

regards, I will keep this issue alive, and I will ask some friends from physics to see what they think.

Thanks a lot for discussing with me