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17.3: Ketone Bodies

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    Kolb et al. BMC Medicine (2021) 19:313 Creative Commons Attribution 4.0 International License,


    Ketone bodies are the name given to two molecules, acetoacetate (a ketone) and D-β-hydroxybutyrate (its reduction product) that derive from the condensation of acetate in the form of acetyl-CoA. Their structures are shown in Figure \(\PageIndex{1}\) below.

    Figure \(\PageIndex{1}\): Structure of ketone bodies

    The are synthesized when acetyl-CoA concentration are high. That would occur under a couple of conditions:

    • fatty acids are being mobilized in adipocytes and broken down into acetyl-CoA through β-oxidation in the mitochondria;
    • the acetyl-CoA produced can not enter the Kreb cycle leading to the buildup of acetyl-CoA in the matrix.

    The latter condition arises when carbohydrate metabolism is compromised so the citric acid cycle has slowed. This occurs if the citric acid cycle intermediate oxaloacetate is being away from the cycle for gluconeogenic synthesis of glucose. Pyruvate, another source of acetyl-CoA (through pyruvate dehydrogenase) is also depleted in gluconeogenesis. Of course we are describing conditions in the liver, the major site of gluconeogenesis and also of ketone body synthesis.

    These conditions are also meet in the underfed or starving state when glycogen supplies are exhausted and fatty acid reserves are being used for energy. Likewise these conditions are met in diabetes when glucose might be abundant in the circulation but its entry into the cell is compromised by problems with insulin signaling.

    Fatty acids released by adipocytes into the blood are bound to serum albumin for transport to tissue. Albumin, with bound fatty acids, does not, however, cross the blood-brain barrier. Ketone bodies are very small and water soluble so they can cross the blood-brain barrier, where they can deliver a soluble equivalent of fatty acids to the brain and other tissues in times of energy need. Ketone bodies can be considered solubilized and easily transportable fatty acid equivalents. They have low pKa values of 3.6 (acetoacetate) and 4.7 (D-β-hydbroxybutyrate) so they can lead to "ketoacidosis" in diabetics. Acetone is nonacidic and sweet-smelling molecule, which can be detected in the breath of diabetics, is produced spontaneously and through catalysis through the decarboxylation of the beta-keto acid acetoacetate. It is also considered a ketone body.

    Blood-Brain barrier

    The blood–brain barrier (BBB) provides protection to the entire nervous system (CNS). Endothelial cells line the surface of all blood vessels. In the CNS their are addition structures that prevent the entry of many molecules. Tight and adherence junctions form between endothelial cells that prevent movement of solutes between the cells. Additional protection is provided by the basement membrane, glial cells and pericytes. In addition, drug efflux transporters (ABC transporters) move xenobiotics (foreign toxic moleculback through the barrier.

    Nonpolar molecules like dioxygen molecules and low molecular weight can cross the BBB by simple diffusion. Other molecules like glucose and amino acids are carried across by transporters. Larger substance can move in through receptor-mediated endocystosis. Albumin, the major carrier of free fatty acids, like other protein, does not readily pass through the barrier, and its drug-macromolecule complex, cannot cross.

    Figure \(\PageIndex{2}\) below shows a representation of the blood-brain barrier.


    Figure \(\PageIndex{2}\): Structure of the blood-brain barrier. Wilhelm et al. Int. J. Mol. Sci. 2013, 14, 1383-1411; doi:10.3390/ijms14011383. Creative Commons Attribution license

    In this section we will explore the synthesis and utilization of ketone bodies. The synthesis pathway is called ketogeneis, and the degradation pathway is called ketolysis. Ketone body production and use are intimately tried to fatty acid metabolism so it is appropriate to discuss them right after fatty acids oxidation. An overview of both ketogenesis and ketolysis is shown in Figure \(\PageIndex{3}\) below.


    Figure \(\PageIndex{3}\): Overview of the ketogenesis (left) and ketolysis (right) pathways. FFA, free fatty acids; mThiolase,
    mitochondrial thiolase; HMGCS2, hydroxy methylglutaryl-CoA synthase; HMGCL, HMG-CoA lyase; BDH1, mitochondrial βOHB dehydrogenase; MCT1/2, monocarboxylate transporter 1 and 2; SCOT, succinyl-CoA:3-oxoacid-CoA transferase; CS, citrate synthase. Kolb et al. BMC Medicine (2021) 19:313 Creative Commons Attribution 4.0 International License.

    Most acetoacetate is converted to its reduction product, β-hydbroxybutyrate, for blood transport. It can spontaneously or through enzymatic catalysis, can be converted to acetone since it is a β-ketoacid with a built-in electron sink that stabilizes the negative charge in the transition state and intermediate. When ketone bodies enter cells, the β-hydbroxybutyrate is converted back to acetoacetate and eventually to acetyl-CoA.

    Ketogenesis: The details

    Figure \(\PageIndex{4}\) shows the chemical structures of all intermediates in the synthesis of ketone bodies.

    Figure \(\PageIndex{4}\): Ketogenesis

    We'll study three of the reactions in ketogenesis in more detail below.

    Reaction 1: Thiolase

    We have presented the mechanism of thiolase in Step 4 of mitochondrial β-oxidation of fatty acid using the enzyme acetyl-CoA acetyltransferase (ACAT1) also called 3-ketoacyl-CoA thiolase. The final step in beta oxidation pathway involves cleavage of the bond between the alpha and beta carbon by CoASH. This step is catalyzed by beta-keto thiolase and is a thiolytic (as opposed to a cleavage by water - a hydrolysis) reaction. The reaction produces one molecule of acetyl CoA and a fatty acyl CoA that is two carbons shorter. The process repeats until the even chain fatty acid is completely converted into acetyl CoA. The activity of the enzyme is reversible and it can also catalyze the Claisen condensation of two acetyl-CoA molecules into acetoacetyl-CoA, which occurs in the first step of ketogenesis. This step makes a C-C bond.

    Reaction 2: HMG-CoA Synthase

    This enzyme catalyzes the condensation of acetoacetyl–CoA (AcAc–CoA) and acetyl–CoA (Ac-CoA) to form 3-hydroxy-3-methylglutaryl (HMG)–CoA, through the formation of a C-C bond. Forming C-C bonds is critical to all of biosynthesis. It's not a simple process. Cofactors are often used such as thiamine pyrophosphate, as we saw with pyruvate dehydrogenase. The reverse process, breaking a C-C bond, is also difficult unless the atoms are activated. One way to activate a carbon atom is to oxidize it through hydroxylation, often using dioxygen in reactions catalyzed by cytochrome P450. Likewise C-C bond formation often occurs through reaction of radicals in homolytic reactions or through rearrangement of key ionic intermediates. Radical reactions can be between a carbon free radical and heme derivatives. Radical reaction involve heme cofactors (as in P450), non-heme iron proteins (where the iron ion can cycle between different oxidation state), flavoproteins (in which the flavin can undergo both 1 and 2 electron redox steps), radical S-adenosylmethionine (SAM) enzymes, and cobalamins.

    HMG-CoA synthase is somewhat unique in that it forms a C-C bond through activating the methyl group of acetyl-cysteine. The acetyl group comes from an acyl-CoA "donor". The enzyme catalyzes the first committed step in the formation of complex isoprenoids (like cholesterol) and of ketone bodies. The product, 3-hydroxy-3-methylglutaryl HMG–CoA, can either be reduced by HMG-CoA reductase to form mevalonate, which leads to cholesterol synthesis, or cleaved by the enzyme HMG-CoA lyase, to produce acetoacetate, a ketone body.

    Since it catalyzes the first step in cholesterol synthesis, the enzyme HMG-CoA reductase has been a primary focus of drug therapy to reduce high levels of serum cholesterol, which is generally associated with cardiovascular disease. Over 200 million people world-wide are on statins, the primary drugs used to block HMG-CoA reductase activity. Luckily, it doesn't affect HMG-CoA lyase. Gram-positive bacteria also requires the mevalonate pathway.

    HMG-CoA synthase catalyzes a bisubstrate reaction that displays ping-pong kinetics, characteristic of a covalent enzyme intermediate. The first substrate binds to the enzyme and transfer an acetyl group to a nucleophilic Cys 111 in the active site to form an acetyl-Cys 111 intermediae. Free CoA departs. Next the second substrate, acetoacetyl-CoA binds, and condense with the acetyl group donated by acetyl-Cys 111. This condensation involves an enolate. A plausible reaction mechanism is shown in Figure \(\PageIndex{5}\) below.


    Figure \(\PageIndex{5}\): Reaction mechanism for HMG-CoA synthase

    Hence there are three parts of the reaction: acetylation/deacetylation, condensation/cleavage with an enolate intermediate and C-C formation and hydrolysis/dehydration. Figure \(\PageIndex{6}\) below shows an interactive iCn3D model of the Staphylococcus aureus HMG-COA Synthase with bound HMG-CoA and acetoacetyl-CoA (1XPK)

    Staphylococcus aureus HMG-COA Synthase with bound HMG-CoA and acetoacetyl-CoA (1XPK).png

    NIH_NCBI_iCn3D_Banner.svg Figure \(\PageIndex{6}\): Staphylococcus aureus HMG-COA Synthase with bound HMG-CoA and acetoacetyl-CoA (1XPK). (Copyright; author via source). Click the image for a popup or use this external link:

    The biologically active unit (homodimer) is shown with each monmer shown in a different color. The A chain (light cyan) has bound HMG-CoA (HMG) while the B chain (light gold) has acetoacetyl-CoA (CAA) bound. The Glu 79, Cys 111 and His 233 in each subunit are shown in CPK sticks and labeled. Note that the Cys 111 is covalent modified in each subunit.

    Figure \(\PageIndex{7}\) below shows an interactive iCn3D model of the human 3-hydroxy-3-methylglutaryl CoA synthase I with bound CoASH (2P8U)

    human 3-hydroxy-3-methylglutaryl CoA synthase I.png

    NIH_NCBI_iCn3D_Banner.svg Figure \(\PageIndex{7}\): Human 3-hydroxy-3-methylglutaryl CoA synthase I (monomer) with bound CoASH (2P8U). (Copyright; author via source). Click the image for a popup or use this external link:

    The active site side chains are numbered differently (Glu 95, Cys 129 and His 264) compared to the S. aurus protein. Only the monomer is shown in this model.

    Reaction 3: HMG-CoA Lyase

    The lyase reaction is a C-C bond cleavage to produce acetyl-CoA and acetoacetate. A gene knockout of this gene in mice is lethal. Figure \(\PageIndex{8}\) below shows a plausible mechanism for the reaction.

    Figure \(\PageIndex{8}\): Mechanism for HMG-CoA lyase of HMG-CoA. (after Fu et al. The Journal of Biological Chemistry, 285, 26341–26349, August 20, 2010)

    Figure \(\PageIndex{9}\) below shows an interactive iCn3D model of the human HMG-CoA lyase with the competitive inhibitor hydroxyglutaryl-CoA (HG-CoA)(3MP3)

    Human HMG-CoA lyase with inhibitor HG-CoA (3MP3).png

    NIH_NCBI_iCn3D_Banner.svg Figure \(\PageIndex{9}\): Human HMG-CoA lyase with inhibitor HG-CoA (3MP3). (Copyright; author via source). Click the image for a popup or use this external link:

    Only one subunit of the homodimer is shown. The key residues involved in catalysis and binding (Arg 41, Asp 42, HIs 233, His 235 and Cys 266) are shown in sticks, CPK colors and labeled. The competitive inhibitor, 3-hydroxyglutaryl-CoA (without the methyl group) is shown in sticks colored CPK. The S-stereoisomer of HMG-CoA or inhibitor binds, not the R-isomer. A water may shuttle protons between Asp 42 and the C3 hydroxyl of HMG-CoA. Arg 41 facilitates the proper enol/keto tautomer of the product mutation on reaction product enolization. Mutation of Arg 41 have large effect on catalysis.

    We won't go into detail on the remaining enzymes since we have encountered variants of NAD+/NADH dehydrogenase reactions before. The acetoacetate decarboxylase just hastens the already spontaneous decarboxylation of the β-keto ester acetoacetate.

    Regulation of Ketogenesis

    Ketogenesis can be upregulated by hormones such as glucagon, cortisol, thyroid hormones, and catecholamines which lead to fatty acid mobilization. Insulin is the primary hormonal regulator of this process.

    Insulin regulates many key enzymes in the ketogenic pathway, and a state of low insulin triggers the process. A low insulin state leads to:

    • Increased free fatty acids (FFAs) arising from decreased inhibition of hormone-sensitive lipase
    • Increased uptake of FFAs into the mitochondria arising from decreased activation of acetyl-CoA carboxylase, decreasing malonyl CoA, which disinhibits Carnitine Palmitoyltransferase 1 (CPT1)
    • Increased production of ketone bodies arising from increased HMG-CoA activity

    Ketolysis: Utilization of ketone bodies

    Figure \(\PageIndex{10}\) shows the chemical structures in the utilization of ketone bodies.

    Figure \(\PageIndex{10}\) shows the chemical structures in the utilization of ketone bodies.

    Since we have encountered reactions similar to these before, we will not discuss their detailed mechanisms.

    Most organs and tissues can use ketone bodies for energy. They are a major source of energy for the brain when glucose is not available. That makes sense given that fatty acids can't cross the blood-brain barrier. The heart prefers fatty acids for energy but can also use ketone bodies. The liver makes but does not use them since the gene for step 2, beta ketoacyl-CoA transferase is not transcribed. D-β-hydroxybutyrate is first converted to the other ketone body, acetoacetate, for the pathway to continue. The net result is the formation of two acetyl-CoAs that can then enter the citric acid cycle for energy production. Acetone is a dead end ketone body and is excreted in the urine or eliminated in the breath.

    Clinical Significance

    An overproduction of ketone bodies through increased ketogenesis can cause ketoacidosis (given the pKas of the ketone bodies). A type is diabetic ketoacidosis (DKA) in Type I and II diabetes which impair glucose use. This leads to liver gluconeogensis and resulting depletionof oxaloacetate and pyruvate, impacting the ability of acetyl-CoA to enter th citric acid cycle.

    On presentation, patients are usually very dehydrated from being hyperglycemic. High glucose levels in the blood cause higher osmotic pressure in the vessels and decresed water reabsorption in the kidneys. They often have accompanying symptons like confusion, nausea, vomiting, and abdominal pain. Because of the acidosis, patients often breathe very deeply and rapidly to eliminate carbon dioxide, which can cause and cause respiratory alkalosis. Ketoacidosis also can occur with severe alcoholism and prolonged starvation.

    Ketogenic Diet

    The ketogenic diet, characterized by high fat and low carbohydrate consumption, can lead to weight loss. It shifts metabolism to more closely resemble the fasting state. It's long term effects are not clear but some have suggested the use of ketone-based drugs to mimic the effect of ketone bodies. Fasting can also lead to ketogenic state and it has been used for the treatment of epilepsy before drug became available. It is important to differentiate this attempt to induced a ketogenic state from diabetic ketoacidosis.

    17.3: Ketone Bodies is shared under a CC BY 4.0 license and was authored, remixed, and/or curated by Henry Jakubowski via source content that was edited to conform to the style and standards of the LibreTexts platform; a detailed edit history is available upon request.