How does krebs cycle produce atp

While these studies show the cytosolic processing of citrate to be generally a pro-inflammatory event, citrate itself has been shown to inhibit the HIF asparaginyl hydroxylase [factor inhibiting HIF FIH ] Increased flux through CIC and the cytosolic processing of citrate has, therefore, been shown to be of importance in the activation of macrophages, DCs, and NK cells. Acetyl-CoA is not only a substrate for de novo lipogenesis, it also is an important cofactor for the acetylation of histones and non-histone proteins Lysine acetylation is reversible and so provides a very useful mechanism for the regulation of gene expression and general protein function Acetyl-CoA cannot travel across cell membranes, and so to exert its effects it must be generated in different cellular compartments The generation of acetyl-CoA from NAA is more commonly associated with processes in the brain and it has also been shown to be a source of nuclear and cytosolic acetyl-CoA in brown adipose tissue ATP-citrate lyase links metabolism to histone acetylation as it converts glucose-derived citrate to acetyl-CoA and it has been found to localize to both nucleus and cytoplasm Citrate is small enough to diffuse across nuclear pores allowing for acetyl-CoA to be produced in either cellular compartment, and siRNA-mediated knockdown of ACLY reduced global histone acetylation Since ACLY is upregulated in lipopolysaccharide LPS -stimulated macrophages 49 , it would be interesting to see where ACLY localized to and if there was a direct effect on the expression of glycolytic genes due to changes in histone acetylation.

ACLY has been shown to control glucose to acetate switch. ACLY-deficient cells upregulate ACSS2 allowing for the production of acetyl-CoA from acetate, ensuring cell viability and providing substrates for both fatty acid synthesis and histone acetylation As previously discussed, in glucose-deprived conditions, increased flux through CIC can sustain NADPH levels in glucose-deprived activated macrophages Acetylation of CIC increases in glucose-deprived growth conditions compared to media containing glucose.

By reconstituting liposomes with mitochondrial extracts, it was shown that the acetylation of CIC causes an increase in V max for citrate Acetate is taken up and processed via the Krebs cycle to produce citrate. Though metabolic reprogramming differs in activated T cells compared to macrophages and DCs, this highlights the importance of the citrate pathway in control of both the metabolism of immune cells and their production of pro-inflammatory mediators, unlike in T cells.

No work has yet been carried out to directly link citrate-derived acetylation in M1 macrophages or DCs, however, histone acetylation is important in macrophage activation and DC differentiation. IL-6 and IL production are both regulated on histone and non-histone protein acetylation, respectively 78 , Therefore, it is likely that acetylation plays a role in the regulation of immune cell metabolism.

Histone acetylation downstream of ACLY has been shown to be of importance in M2 macrophage activation While STAT6 is the major regulator of IL4 induced genes a subset of genes important in the regulation of cellular proliferation and the production of chemokines are under additional control of an Akt—mTORC1 signaling pathway.

Covarrubias et al. They suggest that certain transcription factors and histone acetyltransferases, e. Malonyl-CoA is the cofactor required Lysine-malonylation has been shown to play a role in the regulation of mitochondrial function, FAO and glycolysis 86 , Notably histone malonylation does not occur at the N-terminal tail as happens with acetylation, suggesting that the regulatory role these two modification carry out may be very functionally different to acetylation A large number of proteins involved in fatty acid metabolism are malonylated, including ACLY.

However, no studies have yet been carried out regarding the functional consequence of lysine-malonylation in immune cells. While the accumulation of citrate caused by the IDH1 breakpoint in the TCA cycle can be used to fuel fatty acid synthesis and histone acetylation, another fate of this citrate is the production of itaconate Figure 3.

First identified in as a product of the distillation of citric acid, itaconate has recently become a focus of the field of immunometabolism due to its potential role as an anti-inflammatory modulator. Itaconate is derived from citrate produced in the Krebs cycle and, in M1 macrophages, is one of the most highly induced metabolites following LPS treatment Citrate is acted on by the mitochondrial aconitase 2 ACO2 to produce cis-aconitate.

Cis-aconitate is decarboxlylated by cis-aconitate decarboxylase, also known as immune-responsive gene 1 IRG1 , to produce itaconate. Itaconate has long been used in an industrial setting and is produced on an industrial scale as a fermentation product of Aspergillus terreus for use in the creation of polymer formation Figure 3. Citrate-derived itaconate. Citrate is converted to cis-aconitate by the Krebs cycle enzyme ACO2. Activation of macrophages with LPS causes induction of IRG1 which can produce itaconate by the decarboxylation of cis-aconitate.

Itaconate is toxic to microorganisms expressing ICL, a key component of the glycoxylate shunt in bacteria. In , it was found that immunoresponsive gene 1 Irg1 is highly upregulated in peritoneal macrophages following LPS stimulation 89 , and has since been seen to be upregulated in the blood of human sepsis patients 90 and in the time during embryo implantation 91 , Despite lacking a sequence targeting it, there IRG1 has been found to associate with the mitochondria 93 , It was only in that itaconate was identified in multiple studies in an immune context and in that IRG1 and itaconate were connected Itaconate was seen in the lungs of mice infected with Mycobacterium tuberculosis MTB and was not present in the lungs of control mice In a separate study, itaconate was shown to be secreted by the macrophage cell line RAW Michelucci et al.

They further showed by isotope-labeling that itaconate was derived from citrate. Genetic silencing of Irg1 causes macrophages to lose their bactericidal activity, which was due to decreased amounts of itaconate and the loss of its inhibitory effect on isocitrate lyase ICL , a crucial enzyme of the glycoxylate shunt in bacteria The glycoxylate shunt is a means for bacteria to survive in conditions of low glucose availability where acetate is the primary fuel source.

Succinate enters the Krebs cycle and glycoxylate is then converted to malate by malate synthase. Malate can be processed to oxaloacetate by MDH as in the normal reactions of the Krebs cycle. Some bacteria are able to degrade itaconate, producing acetyl-CoA and pyruvate, due to the expression of genes that encode for itaconate-CoA transferase, itaconyl-CoA hydratase, and S -citramalyl-CoA ligase.

Possession of these genes allows Pseudomonas aeruginosa and Yersinia pestis to survive in activated macrophages There is a discussion as to the relevance of these studies due to differences in concentrations of itaconate used both in terms of the variety of concentrations used exogenously to inhibit bacterial growth and the range in reported intracellular concentrations , It may be that intracellular itaconate is concentrated in vacuoles, and whole cell analysis will not adequately represent this, and that measuring the concentration of secreted itaconate in cell culture media does not determine what the local concentration would be.

While the effect of itaconate on bacterial survival has been well documented, more recent work has sought to elucidate the effect that a high intracellular concentration of itaconate has on the immune cells that produce it. Dimethyl itaconate DMI has been used in several studies as a cell permeable itaconate analog to boost the intracellular levels of itaconate.

The authors suggest that this is due to the ability of itaconate to inhibit SDH, and they showed itaconate to inhibit a purified form of SDH. As SDH also acts as complex II of the ETC, this highlights the ability of endogenous itaconate to regulate mitochondrial metabolism and is consistent with other reports of itaconate competitively inhibiting SDH, albeit weakly, the first of which was in — This led Lampropoulou et al. When succinate accumulates and is oxidized by SDH, it will produce a large amount of coenzyme Q.

IRG1 has also been shown to play a role in the establishment of endotoxin tolerance in LPS-tolerized macrophages. Several other elements regulating IRG1 expression and, therefore, itaconate production have also recently been identified. Inhibition of branched-chain aminotransferase 1 in human monocyte-derived macrophages decreased levels of glycolysis and oxygen consumption while also reduced IRG1 mRNA and protein levels as well as itaconate production A major issue with the study of the functional effect of itaconate in macrophages to date has been the use of DMI.

DMI was utilized as it is cell permeable, however, it has been shown that while DMI boosts the level of itaconate in the cell it is not itself metabolized to itaconate El Azzouny et al. The authors speculate that the effects of DMI on macrophage metabolism may be due to an ability to act as a cysteine alkylating agent or to alter redox homeostasis. They further suggest that, though one has not been identified, it is possible a cell surface receptor for itaconate exists that DMI would be able to bind.

While the effects of studies carried out utilizing DMI have been drawn into question, the body of work carried out using genetic inhibition or deletion of Irg1 and the striking amount by which Irg1 mRNA and itaconate synthesis are upregulated in activated immune cells still leaves it worthy of further investigation. Our understanding of immune cell metabolism has come far since the early observations that activated macrophages were highly glycolytic , It is now well accepted that these pathways play a part outside of their traditional energetic and biosynthetic roles.

The discovery that the Krebs cycle is not complete in activated M1 macrophages and DCs highlights the importance of the withdrawal of citrate from the cycle for DC activation, the production of pro-inflammatory mediators and for the generation of itaconate.

Citrate links many important cellular processes, bridging carbohydrate and fatty acid metabolism and protein modification. Its role in producing acetyl-CoA for the acetylation of histones may turn out to be its most striking role in regulating immune cell function. The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

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Cell 1 — A conserved family of Prolylhydroxylases that modify HIF. Step 8: Malate is oxidized to produce oxaloacetate, the starting compound of the citric acid cycle. Learning Objectives State two other names for the citric acid cycle. Briefly describethe function of the citric acid cycle during aerobic respiration and indicate the reactants and products.

Compare where the citric acid cycle occurs in prokaryotic cells and in eukaryotic cells. State the total number of ATP produced by substrate-level phosphorylation for each acetyl-CoA that enters the citric acid cycle. Summary Aerobic respiration involves four stages: glycolysis, a transition reaction that forms acetyl coenzyme A, the citric acid Krebs cycle, and an electron transport chain and chemiosmosis. The citric acid cycle, also known as the tricarboxylic acid cycle and the Krebs cycle, completes the oxidation of glucose by taking the pyruvates from glycolysis, by way of the transition reaction, and completely breaking them down into CO 2 molecules, H 2 O molecules, and generating additional ATP by oxidative phosphorylation.

This flow of hydrogen ions across the membrane through ATP synthase is called chemiosmosis. Chemiosmosis Figure 4. The result of the reactions is the production of ATP from the energy of the electrons removed from hydrogen atoms. These atoms were originally part of a glucose molecule. At the end of the electron transport system, the electrons are used to reduce an oxygen molecule to oxygen ions. The extra electrons on the oxygen ions attract hydrogen ions protons from the surrounding medium, and water is formed.

The electron transport chain and the production of ATP through chemiosmosis are collectively called oxidative phosphorylation. The number of ATP molecules generated from the catabolism of glucose varies. For example, the number of hydrogen ions that the electron transport chain complexes can pump through the membrane varies between species.

Another source of variance stems from the shuttle of electrons across the mitochondrial membrane. The NADH generated from glycolysis cannot easily enter mitochondria. Another factor that affects the yield of ATP molecules generated from glucose is that intermediate compounds in these pathways are used for other purposes.

Glucose catabolism connects with the pathways that build or break down all other biochemical compounds in cells, and the result is somewhat messier than the ideal situations described thus far. For example, sugars other than glucose are fed into the glycolytic pathway for energy extraction.

Other molecules that would otherwise be used to harvest energy in glycolysis or the citric acid cycle may be removed to form nucleic acids, amino acids, lipids, or other compounds. Overall, in living systems, these pathways of glucose catabolism extract about 34 percent of the energy contained in glucose. What happens when the critical reactions of cellular respiration do not proceed correctly? Mitochondrial diseases are genetic disorders of metabolism.

Mitochondrial disorders can arise from mutations in nuclear or mitochondrial DNA, and they result in the production of less energy than is normal in body cells. Symptoms of mitochondrial diseases can include muscle weakness, lack of coordination, stroke-like episodes, and loss of vision and hearing. Most affected people are diagnosed in childhood, although there are some adult-onset diseases.

Identifying and treating mitochondrial disorders is a specialized medical field. The educational preparation for this profession requires a college education, followed by medical school with a specialization in medical genetics. Medical geneticists can be board certified by the American Board of Medical Genetics and go on to become associated with professional organizations devoted to the study of mitochondrial disease, such as the Mitochondrial Medicine Society and the Society for Inherited Metabolic Disease.

The citric acid cycle is a series of chemical reactions that removes high-energy electrons and uses them in the electron transport chain to generate ATP. One molecule of ATP or an equivalent is produced per each turn of the cycle.