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Uronic acid pathway- Steps and Significance
The uronic acid pathway is an alternative pathway for the oxidation of glucose that does not provide a means of producing ATP, but is utilized for the generation of the activated form of glucuronate, UDP-glucuronate which is mainly used for detoxification of foreign chemicals and for the synthesis of Mucopolysaccharides. This pathway also produces Ascorbic acid in certain animals.
The unutilized Glucuronate produced in this pathway is converted to Xylulose-5 -P which is further metabolized through HMP pathway.
Steps of Uronic acid pathway
The entry of glucose is in the phosphorylated form (Glucose-6-P). The phosphorylation is catalyzed by Hexokinase/ Glucokinase.
1) Formation of UDP glucose
Glucose 6-phosphate is isomerized to glucose- 1-phosphate in a reaction catalyzed by Phosphoglucomutase, which then reacts with uridine triphosphate (UTP) to form uridine diphosphate glucose (UDPGlc) in a reaction catalyzed by UDPGlc Pyro phosphorylase, as occurs in glycogen synthesis. (Figure-1 and 5)
Figure-1- UDP Glucuronic acid (active form) produced from Glucose-6-P is required for the synthesis of mucopolysaccharides, proteoglycans and is also used for the detoxification of foreign compounds.
2) Formation of D- Glucuronic acid
UDPGlc is oxidized at carbon 6 by NAD+-dependent UDPGlc dehydrogenase in a two-step reaction to yield UDP-glucuronate. (Figure 1 and 5)
UDP-glucuronate is the source of glucuronate for reactions involving its incorporation into proteoglycans or for reactions of substrates such as steroid hormones, bilirubin, and a number of drugs that are excreted in urine or bile as glucuronide conjugates. UDP- G is hydrolyzed to form D- Glucuronic acid.
3) Formation of L- Gulonic acid
Glucuronate is first reduced by the NADPH dependent enzyme, Glucuronate reductase to form L- gulonate (Figure-2 and 5)
Synthesis of Ascorbic acid
This pathway is used by plants and some animals for the synthesis of Ascorbic acid.
L- Gulonate is dehydrated in the presence of enzyme, Aldonolactonase to form L-gulono-δ-location, the direct precursor of ascorbate in those animals capable of synthesizing this vitamin, in an NADPH-dependent reaction. Removal of a pair of hydrogen atoms from L-gulono-δ-lactone, under the effect of the enzyme L-gluconolactone oxidase leads to the formation of 2-keto gulono lactone and that is finally converted to L ascorbic acid. (Figure-2)
Figure 2- Synthesis of ascorbic acid from D- glucuronic acid. The enzyme L- gluconolactone oxidase is absent in human beings and in certain animals as shown by the block at the step.
In humans and other primates, as well as guinea pigs, bats, and some birds and fishes, ascorbic acid cannot be synthesized because of the absence of L-gluconolactone oxidase. It is due to a genetic deficiency of this enzyme. It appears that the capacity to synthesize ascorbic acid was lost in these species due to a mutation which was not lethal. These species require vitamin C in the diet. Thus a single enzyme defect in the Uronic acid pathway is responsible for the inefficiency to synthesize ascorbic acid in primates.
4) Fate of L- Gulonate in human beings
The Uronic acid pathway is connected to Pentose phosphate pathway through L-gulonate, (Figure-3 and 5) since the latter can be converted to an intermediate of the Pentose phosphate pathway as follows-
Figure-3- L- Gulonate is oxidatively decarboxylated to form L-xylulose in human beings
5) Fate of L-xylulose
L-Xylulose is converted to the D isomer by an NADPH-dependent reduction to Xylitol, catalyzed by Xylitol dehydrogenase enzymes. The deficiency of Xylitol dehydrogenase causes Essential pentosuria, a clinical state of excess excretion of L- Xylulose in urine.
Xylitol is converted in an NAD+–dependent reaction to form D-Xylulose. The reaction is catalyzed by Xylulose reductase enzyme. D- Xylulose is then phosphorylated to D-Xylulose 5-phosphate and that is metabolized via the pentose phosphate pathway (Figure-4 and 5).
Figure- 4- Fate of L- Xylulose. The flow of electrons is from NADPH to NAD+. Xylulose is phosphorylated by Xylulose kinase to form Xylulose-5- phosphate.
Summary of uronic acid pathway
Figure-5- Uronic acid pathway is a source of glucuronides required for detoxification of drugs as well as hormones and bilirubin. It is also a source of ascorbic acid (not in human beings). Both xylulose as well as ascorbic acid can be metabolized to form oxalates.
Biological significance of Uronic acid pathway- UDP glucuronate the active form of glucuronic acid, can readily donate the glucuronic acid component for the following functions-
1) Detoxification of foreign compounds and drugs– During detoxification, the glucuronate residues are covalently attached to these substances. Since glucuronate residues are strongly polar, their attachment imparts polar character to these substances, making them water soluble and readily excretable. Bilirubin, certain hormones and drugs are made more polar for renal excretion in this manner.
UDP-Glucuronic acid is the Glucuronyl donor, and a variety of glucuronosyl transferases, present in both the endoplasmic reticulum and cytosol, are the catalysts. Molecules such as 2-acetylaminofluorene (a carcinogen), aniline, benzoic acid, meprobamate (a tranquilizer), phenol, and many steroids are excreted as glucuronides. The glucuronide may be attached to oxygen, nitrogen, or sulfur groups of the substrates. Glucuronidation is probably the most frequent conjugation reaction.
Conjugation of Bilirubin-Bilirubin is nonpolar and would persist in cells (e.g., bound to lipids) if not rendered water-soluble. Hepatocytes convert bilirubin to a polar form, which is readily excreted in the bile, by adding Glucuronic acid molecules to it. This process is called conjugation.The conjugation of bilirubin is catalyzed by a specific Glucuronyl transferase. The enzyme is mainly located in the endoplasmic reticulum, uses UDP-Glucuronic acid as the glucuronosyl donor, and is referred to as bilirubin-UGT. Bilirubin Monoglucuronide is an intermediate and is subsequently converted to the diglucuronide. Most of the bilirubin excreted in the bile of mammals is in the form of bilirubin diglucuronide (Figure-6).
Figure-6- Conjugation of bilirubin takes place in the liver. The reaction is catalyzed by UDP glucuronyl transferase. Bilirubin diglucuronide is the water-soluble, conjugated form of bilirubin.
2) Synthesis of Mucopolysaccharides- Glucuronic acid is also required for the synthesis of Mucopolysaccharides such as hyaluronic acid and heparin, which contain glucuronic acid as an essential component.
Clinical Significance of Uronic acid pathway
A) Diminished activity of Bilirubin UDP Glucuronyl Transferase (UGT)
1) Neonatal “Physiologic Jaundice
This transient condition is the most common cause of unconjugated hyperbilirubinemia. It results from an accelerated hemolysis around the time of birth and an immature hepatic system for the uptake, conjugation, and secretion of bilirubin. Not only is the bilirubin-UGT activity reduced, but there probably is reduced synthesis of the substrate for that enzyme, UDP-glucuronic acid. Since the increased amount of bilirubin is unconjugated, it is capable of penetrating the blood-brain barrier when its concentration in plasma exceeds that which can be tightly bound by albumin (20–25 mg/dL). This can result in a hyperbilirubinemic toxic encephalopathy, or kernicterus, which can cause mental retardation. Because of the recognized inducibility of this bilirubin UGT enzyme system, phenobarbital has been administered to jaundiced neonates and is effective in this disorder. In addition, exposure to blue light (phototherapy) promotes the hepatic excretion of unconjugated bilirubin by converting some of the bilirubin to other derivatives such as maleimide fragments and geometric isomers that are excreted in the bile.
2) Crigler-Najjar Syndrome, Type I; Congenital Nonhemolytic Jaundice
Type I Crigler-Najjar syndrome is a rare autosomal recessive disorder. It is characterized by severe congenital jaundice (serum bilirubin usually exceeds 20 mg/dL) due to mutations in the gene encoding bilirubin-UGT activity in hepatic tissues. The disease is often fatal within the first 15 months of life. Children with this condition have been treated with phototherapy, resulting in some reduction in plasma bilirubin levels.
3) Crigler-Najjar Syndrome, Type II
This rare inherited disorder, also results from mutations in the gene encoding bilirubin-UGT, but some activity of the enzyme is retained and the condition has a more benign course than type I. Serum bilirubin concentrations usually do not exceed 20 mg/dL. Patients with this condition can respond to treatment with large doses of phenobarbital.
4) Gilbert Syndrome
Again, this relatively prevalent condition is caused by mutations in the gene encoding bilirubin-UGT. It is more common among males. Approximately 30% of the enzyme’s activity is preserved and the condition is entirely harmless.
B) Essential Pentosuria
Essential Pentosuria is the condition in which an unusual reducing substance, one of the pentose sugars, is constantly excreted in the urine and gives a positive reaction on testing with Benedict’s solution. It is a rare hereditary disease which has been included by Garrod (1923) among the inborn errors of metabolism. Its occurrence was first described in 1892, but since then only about 200 cases have been recorded in the literature, the disorder occurred almost entirely in the Jewish race.
Biochemical defect- The enzyme that causes conversion of L-Xylulose to Xylitol is deficient. As a result the excess of L-Xylulose is excreted in urine.
Clinical Manifestations- It may go unnoticed or it may be a chance finding on routine examination of urine. There are no signs and symptoms associated with it. Various drugs increase the rate at which glucose enters the uronic acid pathway. For example, administration of barbital or chlorobutanol to rats results in a significant increase in the conversion of glucose to glucuronate, L-gulonate, and ascorbate. Aminopyrine and antipyrine increase the excretion of L-xylulose in pentosuric subjects.
Diagnosis-It can be misdiagnosed with renal glycosuria or mild diabetes mellitus. The Qualitative Benedict’s test for reducing substances is given positive in this condition. Bial’s test and fasting blood glucose estimation can rule out renal glycosuria and diabetes mellitus.
The identification of urinary xylulose has been greatly facilitated by the introduction of paper chromatography.
Treatment- No treatment is required for this defect.