Glycolysis exam questions

Exam questions






1.   Which of the following regulates the glycolytic pathway?




a.    Phosphoglycerate mutase


b.   Enolase


c.    Pyruvate kinase


d.   Hexose isomerase


e.    Glyceraldehyde 3-phosphate dehydrogenase




2.   Select the correct multiple answer using ONE of the keys A, B, C, D or E as follows:


A. 1, 2 and 3 are correct


B. 1 and 3 are correct


C. 2 and 4 are correct


D. only 4 is correct


E. all are correct




In the glycolytic pathway, which of the following reactions involve the production of ATP?



1.   Phosphoenolpyruvate –> pyruvate

2.   Fructose 6-phosphate –> fructose 1,6-bisohosphate

3.   1,3-bisphosphoglycerate –> 3-phosphoglycerate

4.   Glyceraldehyde 3-phosphate –> 1,3-bisphosphoglycerate

Defenders of Mitochondria

Alright my Biochem peeps,

Here is my 2nd review article. It is based on the target therapies of cancer treatment. In particular, this article focuses on the drug known as dichloroacetate which targets a mitochondrial enzyme known as pyruvate dehydrogenase kinase. This enzyme inhibits the action of pyruvate dehydrogenase as well as increase the metabolism of cancer cells that facilitate cancer proliferation. This article is quite interesting and thought-provoking. It will aid your understanding of the concepts of glycolysis, Krebs, ETC and oxidative phosphorylation. It is also marvelous to read the mechanism of action of cancer cells and the intricacies of their metabolism that fosters survival and proliferation. Take a read and see for yourself.

Michelakis, Evangelos D. and Gopinath Sutendra. 2013. “Pyruvate dehydrogenase kinase as a novel therapeutic target in oncology.” Frontiers in Oncology 3(38): 1-11. doi: 10.3389/fonc.2013.00038

Pyruvate dehydrogenase kinase as a novel therapeutic target in oncology.

This article is not a study as in the previous posting, but rather delves into the strategies utilized in cancer treatment in particular targeting more efficient, effective and less destructive methods of cancer therapy. This article deals mainly with investigating the effects of inhibition of an enzyme, pyruvate dehydrogenase kinase (PDK) as well as the applications of similar techniques that promote glucose oxidation as a viable option in metabolic targeting of neoplastic cells. The authors also explore the pre-clinical and clinical evidence of the effectiveness of these inhibitors on different cancers.

Neoplastic cells have a unique metabolic property in which there is both molecular and genetic signalling that converts glucose oxidative phosphorylation in the mitochondria of cells to glycolysis in the cytosol even under normal conditions. This results in the increase in glucose consumption as well as lactate production in cancer cells. This process is referred to as the Warburg effect after Otto Warburg who published this hypothesis in 1956. This means that cancer cells depend on glycolysis as its primary energy source as well as the source of macromolecular building blocks through glycolytic intermediates.

This is initiated by the activation of hypoxia-inducible factor 1α (HIF-1α). HIF-1α is activated in hypoxic-like environments and then stimulates the expression of glycolytic enzymes, glucose transporters and mitochondrial enzymes that increase the glycolytic pathway as well as formation of lactate. One important mitochondrial enzyme that is activated is, PDK. PDK  is considered the gatekeeper of the mitochondria since it regulates the flux of pyruvate into the mitochondria to undergo oxidative phosphorylation. PDK does this by inhibition of pyruvate dehydrogenase that performs the link reaction that decarboxylates pyruvate to Acetyl-CoA to enter the Krebs cycle. This inhibition leads to pyruvate not entering the mitochondria and therefore is converted to lactate in the cytosol by lactate dehydrogenase. Oncogenes are responsible for the maintenance of hypoxic state for cancer cells to continue to activate HIF. In cancer cells, there is also a loss of p53 gene expression that was responsible for mitochondrial DNA repair and expression cytochrome c oxidase enzymes that aid in apoptosis of the cell. Loss of p53 directly induces the expression of PDK and increases the stability and transcriptional activity of HIF. Additionally, HIF also increase pro-angiogenic and metastasis-promoting factors such as VEGF and SDF-1.

With increase in glycolytic enzymes, there is an increase in glycolysis as well as these glycolytic enzymes are utilized to aid in anti-apoptotic activities, activation of transcription in neoplastic cells, inhibition of mitochondrial function. Enzymes such include, glyceraldehyde 3-phosphate dehydrogenase and hexokinase. An isoform of hexokinase, hexokinase 2 is up regulated in neoplastic cells and then translocated to the mitochondrial transition pore, a channel responsible for the release of apoptotic factors as well as the regulation of anions the facilitate oxidative phosphorylation. Thus, hexokinase 2 facilitates resistance to apoptosis as well as increase in glycolysis in tumour cells. The glycolytic intermediates may also be funnelled to alternative pathways to produce amino acids, nucleic acids and lipids.

The mechanism of action of cancer cells lies in the alteration of the mitochondria of the cell. The mitochondria are responsible for the intrinsic apoptotic mechanisms of the cells and suppression of mitochondrial function is critical to cancer cell proliferation. One method employed by cancer cells is the alteration of the mitochondrial membrane potential of the inner mitochondrial membrane. This membrane potential is crucial to oxidative phosphorylation and electron transport chain. This membrane potential is akin to the proton motive force that is generated as hydrogen ions are pumped into the inter-membrane space of the mitochondrion during electron transport chain. This membrane potential is also critical to maintenance of intracellular calcium levels since mitochondria act like calcium sinkholes that facilitate the influx of calcium.

Neoplastic cells, inhibit glycogen synthase kinase in the cytosol of the cells that lead to the translocation of hexokinase 2 to the mitochondrial membrane. Hexokinase 2 then binds to the inner membrane and blocks and inhibits the voltage-dependent anion channel found in the mitochondrial transition pore that is responsible for the efflux of anions to maintain membrane potential of the mitochondria that is crucial for oxidative phosphorylation and ETC. This results in the hyperpolarization of the membrane and prevents the opening of the channel in the mitochondrial transition pore that is responsible for release of apoptotic factors such as mitochondrial reactive oxygen species, cytochrome c oxidase  as well as apoptotic inducing factor.

The suppression of mitochondrial functioning leads to the decrease in oxidative phosphorylation as well as the release of apoptotic factors. With diminishing functioning, the mitochondria of tumour cells begin to degenerate and further increase the proliferative potential of the neoplastic cell. However, this information can be utilized in targeting specific cancers especially solid tumours.

A drug known as dichloroacetate is able to penetrate most cell membranes and tissues that are not accessible to chemotherapy treatments such as the brain. This molecule is able to activate pyruvate dehydrogenase by inhibition of pyruvate dehydrogenase kinase and its isoforms. Dichloroacetate carboxylic group forms a salt bridge with Arg154 residue of the active site of pyruvate dehydrogenase kinase. Dicholoroacetate is able to decrease the membrane potential of inner mitochondria and aid in normalization of mitochondrial function in cancer cells. However, it does not affect non-cancer cells. It functions to increase apoptotic activity in the cancer cells as well as to increase the intake of pyruvate into the mitochondria and increase more ETC./oxidative phosphorylation. This decreases the production of lactate and the alternate pathways of glycolytic intermediates.

Subsequently, there is a deactivation of HIF since HIF is highly regulated by the TCA intermediate, alpha ketogluterate. Hence, an increase in TCA cycle leads to increase production of alpha ketogluterate that facilitates the inhibition of the action of HIF. This results in decrease tumour perfusion, glucose uptake, resistance to apoptosis as well as decrease production of macromolecules essential for tumour proliferation. DCA has also shown to increase p53 activity in cancer cells leading increase activation of apoptotic action by the mitochondria.

DCA is therefore very effective at selectively targeting cancer metabolism by inhibition of pyruvate dehydrogenase kinase. Clinical evidence of this has been observed in non-small cell lung carcinoma, breast carcinoma, glioblastoma, colon cancer, prostate and endometrial cancer as well as leukaemia.

However, other therapies may mimic the action of DCA and acquire similar results. Some therapies include the inhibition of lactate dehydrogenase to facilitate the channelling of pyruvate into the mitochondria to undergo oxidative phosphorylation. Similarly, decrease activation of the M2 isoform of pyruvate kinase that is expressed in neoplastic cells may prove to be another selective target for cancer treatment.

In concluding this article, the authors note that the aforementioned techniques are not a permanent solution to cancer therapy since there is no absolute action of these inhibitors on the metabolism of cancer.

My initial thoughts of this paper, was that it was long-winded and boring. However, as I continued past the introduction, the content became more interesting and captivating. The explanations and the organization of the information laid a good base level to which anyone reading was able to easily follow. It was greatly surprised at the intricacies of cancer metabolism and the very dynamic alterations made for the survival and proliferation of these cells. I knew that the mitochondria of cells were integral to the optimal functioning of the cell, nevertheless, I had no knowledge of the specific actions involved in mitochondrial functioning.

The authors aptly summarized the pathophysiology of neoplastic cells and then linked this to the action of the small DCA molecules and its efficiency in cancer treatment through selective inhibition. The importance of pyruvate dehydrogenase kinase was also elucidated. I was also intrigued by the importance of glycolysis to cancer metabolism and how effective glycolytic pathway is to the rapid growth and sustainability of the tumour.

However, it must be stated that even though DCA and the other techniques mentioned in this paper would prove highly valuable to oncology, there is no absolute solution since neoplastic cells have the ability to adapt and evolve to avoid the actions of these target drugs. There is no certainty that these therapies will show significant reduction in tumour proliferation in cancer patients and is subjected to individual differences. These therapies also have the potential to produce effects on non-cancer cells with prolonged use and high doses. Therefore, I am hesitant to believe that DCA and similar inhibitory therapies are a permanent solution to reducing the prevalence of cancer.

Overall, I think that this is a groundbreaking and lifesaving research in oncology since it will prove to reduce the mortality and morbidity of cancer. It is quite exciting to note that this method is a good step forward in finding more advance and effective treatment for a serious medical condition. Clinical evidence thus far has been positive and with more research, the methods of cancer treatment may develop into a simple cure like penicillin.


Parasite and Neoplastic cells Villain

Hey My Biochem peeps,

Hereunder is a review of a new study on the discovery of a new irreversible enzyme inhibitor that may be useful in the treatment of the parasitic infection, Leishmania as well as the treatment of cancers. This inhibitor targets pyruvate kinase, the last enzyme in the glycolytic pathway. Pyruvate kinase is responsible for the production of pyruvate as well as ATP. So I hope you find it as interesting as I did. Enjoy and Happy studying…….

Morgan, Hugh P., Martin J. Walsh, Elizabeth A. Blackburn, Martin A. Wear, Matthew B. Boxer, Min Shen, Henricke Veith, Iain W. McNae, Matthew W. Nowicki, Paul A. M. Michels, Douglas S. Auld, Linda A. Fothergill-Gilmore and Malcolm D. Walkinshaw. 2012. “A new family of covalent inhibitors block nucleotide binding to the active site of pyruvate kinase.” Biochem. J. 448: 67-72. doi:10.1042/BJ20121014.


A new family of covalent inhibitor block nucleotide binding to the active site of pyruvate kinase.

Pyruvate kinase is the last enzyme in the glycolytic pathway and catalyses the substrate-level phosphorylation of phosphoenolpyruvate to pyruvate and converting ADP to ATP in the process.  Human Pyruvate kinase consist of four isozymes: RPYK (Erythrocytes), LPYK (Liver), M1PYK (Muscle), M2PYK (Embryonic Tissues and Neoplastic cancer cells). Most of these isozymes are allosterically regulated by fructose 1, 6-bisphosphate meaning that increase in in fructose 1, 6-bisphosphate will lead to increase activation of pyruvate kinase. M1 isoform is constitutively active.

Leishmania mexicana is a trypanosome parasite that causes Leishmaniasis. It is transmitted by the bite of infected sand flies (Tsetse Fly). This infection results in skin sores, fever, weight loss as well as splenomegaly and hepatomegaly (Medline Plus 2012). Leishmania mexicana possesses similar pyruvate kinases as human however, trypanosome pyruvate kinase (LmPYK) is regulated by fructose 2,6-bisphosphate.

According to Morgan et al., parasitic infections are usually treated by the sulphur drug, Suramin that inhibits 7 out of the 10 glycolytic enzymes of Trypanosoma brucei. This drug works by mimicking ATP/ADP and competitively binds to the active site of pyruvate kinase to inhibit the formation of ADP. It is believed that saccharine inhibitors may have the same effect on LmPYK as suramin. This paper utilizes the saccharine derived compound to inhibit LmPYK activity and hence induce cell death in this parasite. The inhibitor is known as {4-[(1,1 dioxo-1,2-benzothiazol-3-yl)sulfonyl] benzoic acid} or DBS.

Through experimentation, this study showed that DBS was able to bind to the active site of pyruvate kinase, specifically form covalent bonding with the Lys335 residue present in the rim of the active site of pyruvate kinase. This covalent binding leads to the modification of Lys335 that prevents ADP from binding and forming ATP. This process leads to cessation of glycolysis and ATP  generation as well as pyruvate production in the parasite leading to cell death.

First, utilizing purified and characterized wild type and mutant k335R forms of LmPYK, an inhibitor assay was made in order to determine the rate of inhibition of DBS on LmPYK. The inhibitor assay contained a buffer, NADH, lactate dehydrogenase, LmPYK, Phosphoenolpyruvate and inhibitor mix of DBS. A control was created in the same manner except for replacing the inhibitor mixture with buffer solution. The inhibitor assay and control were incubated at 25oC water bath throughout the experiment. ADP was then added to the reaction mixtures and the decrease in absorbance from 340 nm was measured every 20 minutes for 300 mins. This gave the rate of inhibition by DBS.

This experiment illustrated that maximal inhibition was approximately 80% after 250 mins. However, there was never complete inhibition of 100% since after 10 hours, there was heat denaturation of LmPYK due to prolonged incubation in the water bath. The small percentage of pyruvate kinase activity was attributed to the flexibility of the side chains of Lys335. Lys335 residue in the active site has a very flexible R-group side chains that facilitate conformation to allow small access of ADP to the active site of pyruvate kinase to form ATP. However, the affinity of pyruvate kinase to ADP was greatly reduced by the covalent binding of DBS. However, Morgan et al,  posited that 80% inhibition of pyruvate kinase was sufficient to induce cell death in the parasite.

Next, a preparation of an inhibitor-modified LmPYK was made and crystallized in order to identify the structure of the LmPYK-DBS complex. The crystals formed were anisotropic and demonstrated covalent modification of Lys335 residues at the active sites of LmPYK. This modification was located at the adenine-binding site that leads to the blocking of ADP/ATP from binding. The saccharine group of DBS was covalently bonded to the Lys335 residue at the rim of the cleft of the active site. It was also found that the binding conformation of LmPYK-DBS complex mimics that of the drug Suramin.

Finally, using the mutant K335R, the modification of the Lys335 residue on LmPYK was confirmed. This mutant form of LmPYK was similar in action and kinetics of the wild type, however, it possessed a different Lysine residue at its active site and therefore when DBS  was added, there was no inhibition of mutant K335R. This illustrated that DBS is only able to form covalent bonding and irreversible inhibition of pyruvate kinase if Lys335 is present.

The selectivity of DBS for Lys335 was also investigated using rabbit lactate dehydrogenase and firefly luciferase. These are usually coupling enzymes with pyruvate kinase in the respective animals. However, it was found that even though these enzymes share similar characteristics with pyruvate kinase active sites such as lysine residue in the same position of the active site (LDH) or binds to the hydroxyl group of ribose sugar from NAD (Luciferase), DBS was unable to inhibit the action of these enzymes. Hence, DBS is very selective towards Lys335 residues in the active site of pyruvate kinase.

In concluding, Morgan et al., linked the fact that due to the similarity between LmPYK  and certain human PYK isoforms in terms of Lys335 residues at the rim of the active site, DBS would be an effective inhibitor to both human and trypanosome pyruvate kinases. This is especially important for humans since neoplastic tumour cells as well as erythrocyte pyruvate kinases possess these similarities  and thus DBS would provide a good target drug treatment for cancer.

My thoughts on this paper. Firstly, I was elated that it was not very long and the information was presented in a very organized, easily comprehensible manner. Methods, objectives, rationale and findings were clearly outlined and the language was not too technical so that it was difficult to follow the concepts of the paper. The topic was a very stimulating and informative since I am very interested in diseases and novel therapies to stem the prevalence. Leishmaniasis is especially critical since we are susceptible to this parasitic infection given our tropical geography.

Another interesting note, is the fact that isoforms of human pyruvate kinase such as those found in our red blood cells and cancer cells, are similar to those found in this parasite. This nugget of information may prove valuable in the formulation and production of novel cancer therapies. It has been established that neoplastic cells rely on aerobic glycolysis to generate energy and macromolecular building blocks for growth and metastasis of tumours. Therefore, targeting pyruvate kinase is essential to fighting cancers.

The selectivity of DBS, is an added bonus in drug therapies containing this saccharine since it would be able to only target pyruvate kinase that possesses the Lys335residue at the rim of the active site where ADP/ATP binding usually occurs. Thus, DBS will be able to act on specific cells while leaving the normal non-cancer or non-parasitic cells unaffected.

However, this study was the first to demonstrate the effectiveness of DBS and therefore the findings have not been established. There must be more research into DBS and other analogues to determine whether this phenomenon is a solid avenue to follow in terms of therapy. Little is known about the actual effects of DBS on the human cells since this experiment was done by enzyme testing and no live tissue analysis was performed. Therefore, even though DBS seems like a promising compound that may enhance the efficiency and efficacy of target drugs, there has to be more in-depth research into the compound and the implications of its use.

Overall, this paper was very intriguing and interesting. It brought together concepts of glycolysis, irreversible enzyme inhibition and illustrated how the knowledge of biochemical processes may be applied to medical uses. The research is very poignant since parasitic diseases as well as neoplastic ailments are predominant in modern life. This fosters a greater demand for more efficient and effective methods of treatment for disease. The finding of DBS inhibition on the Lys335 residue offers hope and positive connections to increase the efficacy of future medical interventions.

Thus, I believe that this was an enjoyable and educational reading and I am excited to see and hear about the developments in medicine and zoonoses that have arisen out of this novel study. I recommend that you take a read, the reference is listed at the top of the posting. This topic offers a good outlet to link your understanding of the key concepts of glycolysis and enzyme specificity to developing useful things for society.

Until next posting……..


Glycolysis video review

Hey Biochem folks,

Glycolisis (Biochemistry) by Pitanja sa kolokvijuma Pomozimo kolegama


Here is another video that is focused on Glycolysis. This was a long video of just over 50 minutes and the content focused on a medical based knowledge. The lecturer kept focus on the essentials towards the general board exams for medical students and touched on the key aspects of the glycolytic pathway as well as physiological and pathophysiological points to link the biochemistry of glycolysis.

This video was quite informative and interesting, even though there was much content that was not applicable to our course. I felt that the lecturer’s explanation was comprehensive and easily understandable. However, there was little animation and pictures where sometimes there were periods of just looking at the same diagram as he spoke, that was a little monotonous. However, the content was excellent and the explanations were clear and concise. All of the information he mentioned in this video, I came about in my reading and preparation for the glycolysis blog posts and therefore, I found that this video was a well-summarized and integrative approach to the topic of glycolysis.

Even though, there was a lot of skipping through the stages of glycolysis, except for the irreversible stages, there was a good summary of each stage and the focus on the three regulatory enzymes as well as how these enzymes were regulated and the implications of their regulation, was very helpful. The lecturer clearly outlined the process, reasoning behind the regulation of the enzyme and the unique characteristics of the liver that facilitates a more dynamic glycolytic pathway.

The video, however, did not go into any detail on the fates of pyruvate but rather brushed through the two main pathways of producing lactic acid or acetyl-CoA. There was a general summary on lactic acid production and the implications on the human body. However, the applications of this knowledge to athletes and their increased conditioning that may overcome the lactic acid production with extraneous, vigorous exercise, was the highlight of that section. This application created a deeper understanding into the reasons for the physiological changes that occur due to exercise. A fascinating point was the importance of 2, 3-BPG to oxygen dissociation and it importance towards oxygen transport to foetuses in pregnant women.

In summary, this video was informative, interesting and easy to understand. Even though, the content was less applicable to our syllabus, it still provided accurate and well-integrated real life information on glycolysis. The lecturer was well articulated and seemed very knowledgeable in this topic. Overall, the video was good and I recommend that you look at it in your spare time. The major points of this video are summarized below, but I would suggest that you look for yourself.


The video begins with the lecturer talking about how glucose enters the cell. Glucose enters the cell through the GLUT transporters, as he mentioned and these transporters are tissue specific and have different Km values. This means that the affinity for glucose to enter different cells is dependent on different circumstances. As he continues, the lecturer mentions that GLUT 1 and 3 transporters are found in most cells and have a low Km of 1 mmol value, which means that glucose is able to diffuse steadily into cells. However, GLUT 2 are found in the liver and pancreas and have a high Km value and follows 1st order kinetics in that the increase in glucose concentration leads to increase diffusion through these transporters in to the cell. He also mentioned about GLUT 4 transporters that are found in skeletal muscle and adipose tissues that are insulin-dependent and so will facilitate glucose uptake in response to insulin release. Interestingly enough, these GLUT 4 transporters also are increased in muscle cells in response to increased exercise. So one may allude to the fact that increased GLUT 4 transporters in muscles means an increased demand for glucose to generate energy is critical to maintain functioning.

As the lecturer proceeds, he goes into depth of the process of glycolysis with the focus on the three irreversible, regulated steps of glycolysis. He mentions the importance of phosphorylation of glucose and the fact that that hexokinase required Mg ions as a co-factor. He also explains that there is another enzyme, glucokinase in the liver that performs this phosphorylation reaction to convert glucose to glucose 6-phosphate. Glucokinase is hormone-dependant and is able to trap all excess glucose in the blood into hepatocytes in response to insulin release. It was determined that hexokinase is inhibited by glucose 6-phosphate and therefore, when there is an accumulation of glucose 6-phosphate, it prevents hexokinase from converting anymore glucose. Therefore, the excess glucose is funnelled to the liver.

Next, the lecturer explains the importance of PFK-1 and how it is regulated. AMP is able to activate this enzyme and therefore increase its action to produce fructose 1, 6-bisphosphate. This is because, increase levels of AMP may occur when there is increase in exercise and hence there is more utilization of energy and so need to increase the rate of glycolysis to generate energy. ATP and citrate inhibit PFK-1 since these are both indicators of high energy states. When PFK-1 is inhibited, there is decrease in glycolysis and hence will not be able to go on to make fatty acids, which is essential to lipid production. Therefore in the liver, PFK-2 is present and produces fructose 2, 6-bisphosphate. This product over-rides ATP inhibition and forces PFK-1 to continue glycolysis in the liver. It is a very potent activator of PFK-1 to ensure that glycolysis ensues in the liver. This is essential to continue to utilize the high glucose levels in the blood. PFK-2 is driven by insulin.

Glucagon is also essential to the liver since there is inhibition of PFK-1 in the liver and decreases glycolysis and increases gluconeogenesis. This is critical for fasting states and resting states. The lecturer then explains the implications of these processes on persons with hyperglycaemia such as diabetics.

The lecture continues to explain some alternative pathways for dihydroxyacteone phosphate. He then goes on to proceed through the different stages in the energy generation phase of glycolysis. There is little focus on most of the stages in this phase by the lecturer except to note that glyceraldehyde 3-phosphate dehydrogenase generates NADH from the only oxidation reaction in the pathway and performs phosphorylation via an inorganic phosphate. There is a very brief explanation about substrate level phosphorylation. He also notes that in rbc, there is also the conversion of 1, 3-BPG to 2, 3-BPG.

The lecturer then focuses on the last stage of glycolysis catalysed by pyruvate kinase. It is noteworthy that phosphoenolpyruvate is the highest energy compound of the cell. PEP has twice the amount of energy of ATP. There was also discussion on the pyruvate kinase deficiency and the pathophysiology of the haemolytic anaemia that occurs from this deficiency. During this discussion, he explains the importance of glycolysis, NADH and NADPH to erythrocytes and why the blood glucose levels must be maintained at 5mmol.

The video continues to explain the production of lactate and the effects on the body as well as the case with athletes being able to intake high percentage of oxygen and therefore will have increased rates of glycolysis and increased rates of energy production through TCA and ETC. The lecturer then ties this concept into the oxygen transportation of rbc and the release of oxygen from haemoglobin and the importance of 2, 3-BPG. There is an explanation of oxygen saturation curve and the role of 2, 3-BPG. The video ends with the description of the physiology of oxygen transport from mother to foetus in pregnancy.

Hope you will enjoy it as much as I did. Until next posting….


Fates of Pyruvate

In determining the fate of pyruvate after glycolysis, two questions need to be asked;

  1. 1.   Is there oxygen present i.e. aerobic conditions?
  2. 2.   Are there any mitochondria present to facilitate TCA cycle and ETC?


In aerobic conditions, with mitochondria present, the two molecules of pyruvate produced from one molecule of glucose, is then transported to the mitochondria of the cell where it enters the TCA cycle. Via the TCA and ETC, energy is then generated and NAD+ is regenerated in the ETC.

Pyruvate cannot readily enter the TCA cycle and therefore, must be converted to alternative compound. Pyruvate is converted to Acetyl-CoA in the matrix of the mitochondria by the enzyme PYRUVATE DEHYDROGENASE. Pyruvate dehydrogenase is an enzyme complex that consists of three subunits. It also requires five co-factors in order to effectively catalyse the conversion of pyruvate to Acetyl-CoA. These 5 co-factors are Co-A, TPP, lipoate, FAD and NAD+. During this process, there is loss of a carbon atom from pyruvate to form carbon dioxide. Additionally, NAD+ is reduced to NADH to facilitate the oxidation of pyruvate to acetyl-CoA. Hence, this reaction is an OXIDIZING DECARBOXYLATION.

At the end of the reaction, 2 molecules of pyruvate form 2 molecules of Acetyl Co-A + 2 molecules of carbon dioxide + NADH. This reaction is also IRREVERSIBLE. Once Acetyl CoA is formed, it is combined to oxaloacetate in the matrix of the mitochondria to form citrate in the TCA cycle. This occurs in plant, animal and some microbial cells with the essential requirement that aerobic conditions exist and mitochondria are present.

Pyruvate may also be converted to oxaloacetate, which also enters the TCA cycle to generate energy. This reaction is catalysed by the enzyme, PYRUVATE CARBOXYLASE that is dependent on biotin. Pyruvate undergoes a carboxylation reaction to form oxaloacetate. This is an essential process to replenish the intermediates of the TCA cycle and provide substrates for gluconeogenesis


In anaerobic conditions and in the absence of mitochondria, Fermentation reactions occur. Fermentation may occur by two methods, which result in either the formation of ethanol or lactic acid. The main purpose of fermentation is to regenerate the limited supply of NAD+ in the cell to continue reactions including glycolysis

In yeast cells, fermentation occurs in two steps. 2 molecules of pyruvate are converted to two molecules of ethanol and 2 molecules of carbon dioxide. Firstly, pyruvate is converted to acetaldehyde by the enzyme, PYRUVATE DECARBOXYLASE.  This enzyme utilizes similar co factors to pyruvate dehydrogenase such as TPP, but may also utilize magnesium ions. In this decarboxylation, there is the release of carbon dioxide. This is an IRREVERSIBLE REACTION. Acetaldehyde is then converted to ethanol via ALCOHOL DEHYDROGENASE. During this process, NADH is oxidized to NAD+ and thus, performs a REDOX reaction. This reaction is reversible, however.

Funny enough, alcohol dehydrogenase is found in hepatocytes and aid in the metabolism of ethanol/alcohol after consumption. This enzyme also exists as isoforms, which is mutated in Asians and people of Asian ethnicity. This means that there is defective functioning of alcohol dehydrogenase and thus these people cannot metabolize alcohol quickly and suffer the toxic effects of alcohol quickly.


Fermentation may also occur in human cells under certain conditions. During vigorous exercising, skeletal muscles are unable to obtain an adequate supply of oxygen and therefore there are states of anoxia where the muscles have to undergo anaerobic respiration. In these circumstances, skeletal muscle undergoes fermentation to convert pyruvate to lactate. This process is essential to replenish the supply of NAD+ in order to ensure that glycolysis continues. However, lactic acid is toxic to cells since it causes a change in the pH and leads to acidosis.

In this fermentation reaction, pyruvate is converted to lactate by the enzyme, LACTATE DEHYDROGENASE. This is a slightly reversible reaction. During this process, NADH is oxidized to NAD+, which facilitates the reduction of pyruvate to lactate. Fermentation to form lactic acid also occurs in erythrocytes. Red blood cells lack mitochondria and therefore is unable to carry out TCA cycle and ETC, thus it relies on glycolysis for energy generation. This means that lactate is produced to ensure the quick regeneration of NAD+ to continue the glycolytic pathway.