Pyruvate carboxylase
Encyclopedia
Pyruvate carboxylase is an enzyme
Enzyme
Enzymes are proteins that catalyze chemical reactions. In enzymatic reactions, the molecules at the beginning of the process, called substrates, are converted into different molecules, called products. Almost all chemical reactions in a biological cell need enzymes in order to occur at rates...

 of the ligase
Ligase
In biochemistry, ligase is an enzyme that can catalyse the joining of two large molecules by forming a new chemical bond, usually with accompanying hydrolysis of a small chemical group dependent to one of the larger molecules...

 class that catalyzes the (depending on the species) irreversible carboxylation
Carboxylation
Carboxylation in chemistry is a chemical reaction in which a carboxylic acid group is introduced in a substrate. The opposite reaction is decarboxylation.-Carboxylation in organic chemistry:In organic chemistry many different protocols exist for carboxylation...

 of pyruvate to form oxaloacetate (OAA).
It is an important anaplerotic reaction
Anaplerotic reactions
Anaplerotic reactions are those that form intermediates of a metabolic pathway. Examples of such are found in the Tricarboxylic acid Cycle...

 that creates oxaloacetate from pyruvate. The enzyme is a mitochondrial protein
Protein
Proteins are biochemical compounds consisting of one or more polypeptides typically folded into a globular or fibrous form, facilitating a biological function. A polypeptide is a single linear polymer chain of amino acids bonded together by peptide bonds between the carboxyl and amino groups of...

 containing a biotin
Biotin
Biotin, also known as Vitamin H or Coenzyme R, is a water-soluble B-complex vitamin discovered by Bateman in 1916. It is composed of a ureido ring fused with a tetrahydrothiophene ring. A valeric acid substituent is attached to one of the carbon atoms of the tetrahydrothiophene ring...

 prosthetic group, requiring magnesium
Magnesium
Magnesium is a chemical element with the symbol Mg, atomic number 12, and common oxidation number +2. It is an alkaline earth metal and the eighth most abundant element in the Earth's crust and ninth in the known universe as a whole...

 or manganese
Manganese
Manganese is a chemical element, designated by the symbol Mn. It has the atomic number 25. It is found as a free element in nature , and in many minerals...

 and acetyl CoA, and occurs in liver
Liver
The liver is a vital organ present in vertebrates and some other animals. It has a wide range of functions, including detoxification, protein synthesis, and production of biochemicals necessary for digestion...

 but not in muscle
Muscle
Muscle is a contractile tissue of animals and is derived from the mesodermal layer of embryonic germ cells. Muscle cells contain contractile filaments that move past each other and change the size of the cell. They are classified as skeletal, cardiac, or smooth muscles. Their function is to...

.

Pyruvate carboxylase was first discovered in 1959 at Western Reserve University by M. F. Utter and D. B. Keech. Since then it has been found in a wide variety of prokaryotes and eukaryotes including fungi, bacteria, plants, and animals. In mammals, PC plays a crucial role in gluconeogenesis and lipogenesis, in the biosynthesis of neurotransmitter
Neurotransmitter
Neurotransmitters are endogenous chemicals that transmit signals from a neuron to a target cell across a synapse. Neurotransmitters are packaged into synaptic vesicles clustered beneath the membrane on the presynaptic side of a synapse, and are released into the synaptic cleft, where they bind to...

s, and in glucose-induced insulin secretion by pancreatic islets. Oxaloacetate produced by PC is an important intermediate, which is used in these biosynthetic pathways. In mammals, PC is expressed in a tissue-specific manner, with its activity found to be highest in the liver and kidney (gluconeogenic tissues), in adipose tissue and lactating mammary gland (lipogenic tissues), and in pancreatic islets. Activity is moderate in brain, heart and adrenal gland, and least in white blood cells and skin fibroblasts.

Structure

Structural studies of PC have been conducted by electron microscopy, by limited proteolysis
Proteolysis
Proteolysis is the directed degradation of proteins by cellular enzymes called proteases or by intramolecular digestion.-Purposes:Proteolysis is used by the cell for several purposes...

, and by cloning and sequencing of genes and cDNA encoding the enzyme. Most well characterized forms of active PC consist of four identical subunits arranged in a tetrahedron-like structure. Each subunit contains a single biotin
Biotin
Biotin, also known as Vitamin H or Coenzyme R, is a water-soluble B-complex vitamin discovered by Bateman in 1916. It is composed of a ureido ring fused with a tetrahydrothiophene ring. A valeric acid substituent is attached to one of the carbon atoms of the tetrahydrothiophene ring...

 moiety acting as a swinging arm to transport carbon dioxide
Carbon dioxide
Carbon dioxide is a naturally occurring chemical compound composed of two oxygen atoms covalently bonded to a single carbon atom...

 to the catalytic site that is formed at the interface between adjacent monomers. Each subunit of the functional tetramer contains four domains: the biotin carboxylation (BC) domain, the transcarboxylation (CT) domain, the biotin carboxyl carrier (BCCP) domain and the recently termed PC tetramerization (PT) domain. From the two most complete crystal structures available, an asymmetric and symmetric form of the protein have been visualized. The Staphylococcus aureus
Staphylococcus aureus
Staphylococcus aureus is a facultative anaerobic Gram-positive coccal bacterium. It is frequently found as part of the normal skin flora on the skin and nasal passages. It is estimated that 20% of the human population are long-term carriers of S. aureus. S. aureus is the most common species of...

tetramer in complex with the activator Coenzyme A is highly symmetric, possessing 222 symmetry, and has been confirmed by Cryo-EM studies. In contrast the Rhizobium etli
Rhizobium
Rhizobium is a genus of Gram-negative soil bacteria that fix nitrogen. Rhizobium forms an endosymbiotic nitrogen fixing association with roots of legumes and Parasponia....

, tetramer in complex with ethyl-CoA, a non-hydrolyzable analog of Acetyl-CoA
Acetyl-CoA
Acetyl coenzyme A or acetyl-CoA is an important molecule in metabolism, used in many biochemical reactions. Its main function is to convey the carbon atoms within the acetyl group to the citric acid cycle to be oxidized for energy production. In chemical structure, acetyl-CoA is the thioester...

, possesses only one line of symmetry.
Pyruvate carboxylase uses a covalently attached Biotin cofactor which is used to catalyze the ATP– dependent carboxylation of pyruvate to oxaloacetate in two steps. Biotin is initially carboxylated at the BC active site by ATP and bicarbonate. The carboxyl group is subsequently transferred by carboxybiotin to a second active site in the CT domain, where pyruvate is carboxylated to generate oxaloacetate. The BCCP domain transfers the tethered cofactor between the two remote active sites. The allosteric binding site in PC offers a target for modifiers of activity that may be useful in the treatment of obesity or type II diabetes, and the mechanistic insights gained from the complete structural description of RePC (R. etli) permit detailed investigations into the individual catalytic and regulatory sites of the enzyme.

Reaction mechanism

The reaction mechanism can be subdivided into two partial reactions (see figure to the right). In the first reaction, ATP
Adenosine triphosphate
Adenosine-5'-triphosphate is a multifunctional nucleoside triphosphate used in cells as a coenzyme. It is often called the "molecular unit of currency" of intracellular energy transfer. ATP transports chemical energy within cells for metabolism...

 is carboxylated to produce carbonic phosphoric anhydride [O(O)P(=O)O–C(=O)O] which in turn carboxylates a biotin cofactor that is covalently attached to a lysine residue of the BCCP domain. Carbonic phosphoric anhydride decomposes into carbon dioxide and phosphate prior to attack by the enzyme linked biotin molecule. In most species, this reaction requires acetyl-CoA
Acetyl-CoA
Acetyl coenzyme A or acetyl-CoA is an important molecule in metabolism, used in many biochemical reactions. Its main function is to convey the carbon atoms within the acetyl group to the citric acid cycle to be oxidized for energy production. In chemical structure, acetyl-CoA is the thioester...

 as an allosteric activator binding to the PT domain. In the second reaction, occurring in the CT domain of an adjacent monomer, carbon dioxide is transferred to the acceptor molecule, pyruvate, to form oxaloacetate. The reaction proceeds via the removal of a proton from pyruvate, by an as yet unidentified active site residue, to generate an enol
Enol
Enols are alkenes with a hydroxyl group affixed to one of the carbon atoms composing the double bond. Alkenes with a hydroxyl group on both sides of the double bond are called enediols. Deprotonated anions of enols are called enolates...

ate intermediate. The enolate intermediate than attacks CO2 transiently released from the enzyme linked biotin molecule. The resultant oxaloacetate is released. The biotin molecule is protonated by the aforementioned active site residue and released from the active site of the CT domain to be recarboxylated. The major regulator of enzyme activity, acetyl-CoA, stimulates the cleavage of ATP in the first partial reaction and also it has been shown to induce a conformational change in the tetrameric structure of the enzyme.

Role in gluconeogenesis

During gluconeogenesis
Gluconeogenesis
Gluconeogenesis is a metabolic pathway that results in the generation of glucose from non-carbohydrate carbon substrates such as lactate, glycerol, and glucogenic amino acids....

, pyruvate carboxylase is involved in the synthesis of phosphoenolpyruvate
Phosphoenolpyruvate
Phosphoenolpyruvic acid , or phosphoenolpyruvate as the anion, is an important chemical compound in biochemistry. It has the high-energy phosphate bond found in living organisms, and is involved in glycolysis and gluconeogenesis...

 (PEP) from pyruvate. Pyruvate is first converted by pyruvate carboxylase to oxaloacetate (OAA) in the mitochondrion requiring hydrolysis
Hydrolysis
Hydrolysis is a chemical reaction during which molecules of water are split into hydrogen cations and hydroxide anions in the process of a chemical mechanism. It is the type of reaction that is used to break down certain polymers, especially those made by condensation polymerization...

 of one molecule of ATP
Adenosine triphosphate
Adenosine-5'-triphosphate is a multifunctional nucleoside triphosphate used in cells as a coenzyme. It is often called the "molecular unit of currency" of intracellular energy transfer. ATP transports chemical energy within cells for metabolism...

. The OAA is then decarboxylated and simultaneously phosphorylated, which is catalyzed by one of two isoforms of phosphoenolpyruvate carboxykinase
Phosphoenolpyruvate carboxykinase
Phosphoenolpyruvate carboxykinase is an enzyme in the lyase family used in the metabolic pathway of gluconeogenesis. It converts oxaloacetate into phosphoenolpyruvate and carbon dioxide.It is found in two forms, cytosolic and mitochondrial....

 (PEPCK) either in the cytosol
Cytosol
The cytosol or intracellular fluid is the liquid found inside cells, that is separated into compartments by membranes. For example, the mitochondrial matrix separates the mitochondrion into compartments....

 or in the mitochondria to produce PEP. Under ordinary gluconeogenic condition, OAA is converted into PEP by mitochondrial PEPCK; the resultant PEP is than transported out of the mitochondria via the Citric acid cycle
Citric acid cycle
The citric acid cycle — also known as the tricarboxylic acid cycle , the Krebs cycle, or the Szent-Györgyi-Krebs cycle — is a series of chemical reactions which is used by all aerobic living organisms to generate energy through the oxidization of acetate derived from carbohydrates, fats and...

 carrier system, and converted into glucose by cytosolic gluconeogenic enzymes. However, during starvation when cytostolic NADH concentration is low and mitochrondrial NADH levels are high oxaloacetate can be used as a shuttle of reducing equivalents. As such OAA is converted into malate
Malic acid
Malic acid is an organic compound with the formula HO2CCH2CHOHCO2H. It is a dicarboxylic acid which is made by all living organisms, contributes to the pleasantly sour taste of fruits, and is used as a food additive. Malic acid has two stereoisomeric forms , though only the L-isomer exists...

 by mitochondrial Malate dehydrogenase
Malate dehydrogenase
Malate dehydrogenase is an enzyme in the citric acid cycle that catalyzes the conversion of malate into oxaloacetate and vice versa...

 (MDH). After export into the cytosol, malate is converted back into OAA, with contaminant reduction of NAD+; OAA is subsequently converted to PEP which is available for gluconeogenesis in the cytosol along with the transported reducing equivalent NADH.

Very high levels of PC activity, together with high activities of other gluconeogenic enzymes including PEPCK, fructose-1,6-bisphosphatase and glucose-6-phosphatase in liver and kidney cortex, suggest that a primary role of PC is to participate in gluconeogenesis in these organs. During fasting or starvation when endogenous glucose is required for certain tissues (brain, white blood cells and kidney medulla), expression of PC and other gluconeogenic enzymes is elevated. In rats and mice, alteration of nutrition status has been shown to affect hepatic PC activity. Fasting promotes hepatic glucose production sustained by an increased pyruvate flux, and increases in PC activity and protein concentration; Diabetes similarly increases gluconeogenesis through enhanced uptake of substrate and increased flux through liver PC in mice and rats Similarly to other gluconeogenic enzymes, PC is positively regulated by glucagon
Glucagon
Glucagon, a hormone secreted by the pancreas, raises blood glucose levels. Its effect is opposite that of insulin, which lowers blood glucose levels. The pancreas releases glucagon when blood sugar levels fall too low. Glucagon causes the liver to convert stored glycogen into glucose, which is...

 and glucocorticoid
Glucocorticoid
Glucocorticoids are a class of steroid hormones that bind to the glucocorticoid receptor , which is present in almost every vertebrate animal cell...

s while negatively regulated by insulin
Insulin
Insulin is a hormone central to regulating carbohydrate and fat metabolism in the body. Insulin causes cells in the liver, muscle, and fat tissue to take up glucose from the blood, storing it as glycogen in the liver and muscle....

. Further supporting the key role of PC in gluconeogenesis, in dairy cattle, which have hexose
Hexose
In organic chemistry, a hexose is a monosaccharide with six carbon atoms, having the chemical formula C6H12O6. Hexoses are classified by functional group, with aldohexoses having an aldehyde at position 1, and ketohexoses having a ketone at position 2....

 absorption ability at adequate nutrition levels, PC and the associated gluconeogenic enzyme PEPCK are markedly elevated during the transition to lactation in proposed support of lactose synthesis for milk production.

Aside from the role of PC in gluconeogenesis, PC serves an anaplerotic
Anaplerotic reactions
Anaplerotic reactions are those that form intermediates of a metabolic pathway. Examples of such are found in the Tricarboxylic acid Cycle...

 role (an enzyme catalyzed reaction that can replenish the supply of intermediates in the citric acid cycle) for the tricarboxylic acid cycle
Citric acid cycle
The citric acid cycle — also known as the tricarboxylic acid cycle , the Krebs cycle, or the Szent-Györgyi-Krebs cycle — is a series of chemical reactions which is used by all aerobic living organisms to generate energy through the oxidization of acetate derived from carbohydrates, fats and...

 (essential to provide oxaloacetate), when intermediates are removed for different biosynthetic purposes.

Clinical significance

As a crossroad between carbohydrate and lipid
Lipid metabolism
Lipid metabolism refers to the processes that involve the intercourse and degradation of lipids.The types of lipids involved include:* Bile salts* Cholesterols* Eicosanoids* Glycolipids* Ketone bodies* Fatty acids - see also fatty acid metabolism...

 metabolism, pyruvate carboxylase expression in gluconeogenic tissues, adipose tissues and pancreatic islets most be coordinated. In conditions of over nutrition, PC levels are increased in pancreatic β-cells to increase pyruvate cycling
Pyruvate cycling
Pyruvate cycling commonly refers to an intracellular loop of spatial movements and chemical transformations involving pyruvate. Spatial movements occur between mitochondria and cytosol and chemical transformations create various Krebs cycle intermediates. In all variants, pyruvate is imported into...

 in response to chronically elevated levels of glucose
Glucose
Glucose is a simple sugar and an important carbohydrate in biology. Cells use it as the primary source of energy and a metabolic intermediate...

. In contrast, PC enzyme levels in the liver are decreased by insulin
Insulin
Insulin is a hormone central to regulating carbohydrate and fat metabolism in the body. Insulin causes cells in the liver, muscle, and fat tissue to take up glucose from the blood, storing it as glycogen in the liver and muscle....

; during periods of overnutrition adipocyte tissue is expanded with extreme expression of PC and other lipogenic enzymes. Hepatic control of glucose levels is still regulated in an over nutrition situation, but in obesity induced type 2 diabetes the regulation of peripheral glucose levels is no longer under regulation of insulin.
In type 2 diabetic
Diabetes mellitus type 2
Diabetes mellitus type 2formerly non-insulin-dependent diabetes mellitus or adult-onset diabetesis a metabolic disorder that is characterized by high blood glucose in the context of insulin resistance and relative insulin deficiency. Diabetes is often initially managed by increasing exercise and...

 rats, chronic exposure of β-cells
Beta cell
Beta cells are a type of cell in the pancreas located in the so-called islets of Langerhans. They make up 65-80% of the cells in the islets.-Function:...

 to glucose due to peripheral insulin resistance results in decreased PC enzyme activity and decreased pyruvate cycling The continued overproduction of glucose by hepatocytes causes dramatic alteration of β-cell gene expression with large increases in normally suppressed genes, and equivalent decreases in expression of mRNA for insulin, ion pumps necessary for insulin secretion, and metabolic enzymes related to insulin secretion, including pyruvate carboxylase Concurrently adipose tissue develops insulin resistance causing accumulation of triaglycerols and non-esterified fatty acids in circulation; these not only further impairing β-cell function, but also further decreasing PC expression. These changes result in the decline of the β-cell phenotype in decompensated diabetes.

A deficiency of pyruvate carboxylase
Pyruvate carboxylase deficiency
Pyruvate carboxylase deficiency is an inherited disorder that causes lactic acid and other potentially toxic compounds to accumulate in the blood. High levels of these substances can damage the body's organs and tissues, particularly in the nervous system. Pyruvate carboxylase deficiency is a rare...

 can cause lactic acidosis
Lactic acidosis
Lactic acidosis is a physiological condition characterized by low pH in body tissues and blood accompanied by the buildup of lactate especially D-lactate, and is considered a distinct form of metabolic acidosis. The condition typically occurs when cells receive too little oxygen , for example...

 as a result of lactate
Lactic acid
Lactic acid, also known as milk acid, is a chemical compound that plays a role in various biochemical processes and was first isolated in 1780 by the Swedish chemist Carl Wilhelm Scheele. Lactic acid is a carboxylic acid with the chemical formula C3H6O3...

 build up. Normally, excess pyruvate is shunted into gluconeogenesis
Gluconeogenesis
Gluconeogenesis is a metabolic pathway that results in the generation of glucose from non-carbohydrate carbon substrates such as lactate, glycerol, and glucogenic amino acids....

 via conversion of pyruvate into oxaloacetate, but because of the enzyme deficiency, excess pyruvate is converted into lactate instead. As a key role of gluconeogenesis is in the maintenance of blood sugar
Blood sugar
The blood sugar concentration or blood glucose level is the amount of glucose present in the blood of a human or animal. Normally in mammals, the body maintains the blood glucose level at a reference range between about 3.6 and 5.8 mM , or 64.8 and 104.4 mg/dL...

, deficiency of pyruvate carboxylase can also lead to hypoglycemia
Hypoglycemia
Hypoglycemia or hypoglycæmia is the medical term for a state produced by a lower than normal level of blood glucose. The term literally means "under-sweet blood"...

.

External links


See also

  • Pyruvate carboxylase deficiency
    Pyruvate carboxylase deficiency
    Pyruvate carboxylase deficiency is an inherited disorder that causes lactic acid and other potentially toxic compounds to accumulate in the blood. High levels of these substances can damage the body's organs and tissues, particularly in the nervous system. Pyruvate carboxylase deficiency is a rare...

The source of this article is wikipedia, the free encyclopedia.  The text of this article is licensed under the GFDL.
 
x
OK