Nucleotide metabolism (r ) 1

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Nucleotide metabolism (r ) 1

  1. 1. Dr Falih JaborDr Falih Jabor
  2. 2. Reference BooksReference Books  Lippincotts illustrated review of BiochemistryLippincotts illustrated review of Biochemistry by Champe and Harvey.by Champe and Harvey.  Medical Biochemistry by Baynes andMedical Biochemistry by Baynes and Dominiczak.Dominiczak.
  3. 3. Nucleotide MetabolismNucleotide Metabolism
  4. 4.  IntroductionIntroduction Purine Nucleotide BiosynthesisPurine Nucleotide Biosynthesis Regulation of Purine SynthesisRegulation of Purine Synthesis Catabolism and Salvage of Purine NucleotidesCatabolism and Salvage of Purine Nucleotides Clinical Significances of Purine MetabolismClinical Significances of Purine Metabolism Synthesis of Pyrimidine NucleotidesSynthesis of Pyrimidine Nucleotides Regulation of Pyrimidine SynthesisRegulation of Pyrimidine Synthesis Catabolism and Salvage of Pyrimidine NucleotidesCatabolism and Salvage of Pyrimidine Nucleotides Clinical Significances of Pyrimidine MetabolismClinical Significances of Pyrimidine Metabolism Formation of DeoxyribonucleotidesFormation of Deoxyribonucleotides Interconversion of the NucleotidesInterconversion of the Nucleotides
  5. 5. Ribonucleosides and deoxyribonucleoside phosphates (nucleotides) areRibonucleosides and deoxyribonucleoside phosphates (nucleotides) are essential for all cells. Without them, neither DNA nor RNA can beessential for all cells. Without them, neither DNA nor RNA can be produced and, therefore, proteins cannot be synthesized or cellsproduced and, therefore, proteins cannot be synthesized or cells proliferate.proliferate. 1.1. Nucleotides serve as carriers of activated intermediates in theNucleotides serve as carriers of activated intermediates in the synthesis of some carbohydrates, lipids, and proteins.synthesis of some carbohydrates, lipids, and proteins. 2. They are structural components of several essential coenzymes, for2. They are structural components of several essential coenzymes, for example, coenzyme A,FAD.example, coenzyme A,FAD. 3. Nucleotides, such as cyclic AMP (cAMP) and cyclic GMP (cGMP), serve3. Nucleotides, such as cyclic AMP (cAMP) and cyclic GMP (cGMP), serve as second messengers in signal transduction pathways.as second messengers in signal transduction pathways. 4. Nucleotides play an important role as "energy currency" in the cell.4. Nucleotides play an important role as "energy currency" in the cell. 5. Nucleotides are important regulatory compounds for many of the5. Nucleotides are important regulatory compounds for many of the pathways of intermediary metabolism, inhibiting or activating keypathways of intermediary metabolism, inhibiting or activating key enzymes.enzymes.
  6. 6.  The metabolic requirements for the nucleotides and their bases can beThe metabolic requirements for the nucleotides and their bases can be met by both dietary intake or synthesis de novo from low molecularmet by both dietary intake or synthesis de novo from low molecular weight precursors. The salvage pathways are a major source ofweight precursors. The salvage pathways are a major source of nucleotides for synthesis of DNA, RNA and enzyme co-factors.nucleotides for synthesis of DNA, RNA and enzyme co-factors. Extracellular hydrolysis of ingested nucleic acids occurs through theExtracellular hydrolysis of ingested nucleic acids occurs through the actions of endonucleases, phosphodiesterases and nucleosideactions of endonucleases, phosphodiesterases and nucleoside phosphorylases. Endonucleases degrade DNA and RNA leading to thephosphorylases. Endonucleases degrade DNA and RNA leading to the production of oligonucleotides. Oligonucleotides are further digestedproduction of oligonucleotides. Oligonucleotides are further digested by phosphodiesterases yielding free nucleosides. The bases areby phosphodiesterases yielding free nucleosides. The bases are hydrolyzed from nucleosides by the action of phosphorylases thathydrolyzed from nucleosides by the action of phosphorylases that yield ribose-1-P and free bases. If the nucleosides and/or bases are notyield ribose-1-P and free bases. If the nucleosides and/or bases are not re-utilized the purine bases are further degraded to uric acid and there-utilized the purine bases are further degraded to uric acid and the pyrimidines to beta-alanine and beta-aminoisobutyrate, NH3 and CO2.pyrimidines to beta-alanine and beta-aminoisobutyrate, NH3 and CO2. Both the salvage and de novo synthesis pathways of purine andBoth the salvage and de novo synthesis pathways of purine and pyrimidine biosynthesis lead to production of nucleoside-5'-pyrimidine biosynthesis lead to production of nucleoside-5'- phosphates through the utilization an activated sugar intermediate andphosphates through the utilization an activated sugar intermediate and a class of enzymes called phosphribosyltransferases. The activateda class of enzymes called phosphribosyltransferases. The activated sugar used is 5-phospho--D-ribosyl-1-pyrophosphate, PRPP: PRPP issugar used is 5-phospho--D-ribosyl-1-pyrophosphate, PRPP: PRPP is generated by the action of PRPP synthetase and requires energy in thegenerated by the action of PRPP synthetase and requires energy in the form of ATP as shown:form of ATP as shown: ribose-5-phosphate + ATP -------> PRPP + AMPribose-5-phosphate + ATP -------> PRPP + AMP
  7. 7. Purine Nucleotide BiosynthesisPurine Nucleotide Biosynthesis The major site of purine synthesis is in the liver. Synthesis of the purineThe major site of purine synthesis is in the liver. Synthesis of the purine nucleotides begins with PRPP and leads to the first fully formed nucleotide,nucleotides begins with PRPP and leads to the first fully formed nucleotide, inosine 5'-monophosphate (IMP).inosine 5'-monophosphate (IMP). Inosine monophosphate, IMP begins with 5-phospho-a-ribosyl-1-Inosine monophosphate, IMP begins with 5-phospho-a-ribosyl-1- pyrophosphate, PRPP. Through a series of reactions utilizing ATP,pyrophosphate, PRPP. Through a series of reactions utilizing ATP, tetrahydrofolate (THF) derivatives, glutamine, glycine and aspartate thistetrahydrofolate (THF) derivatives, glutamine, glycine and aspartate this pathway yields IMP. The two indicated enzymes (A and B) are those catalyzingpathway yields IMP. The two indicated enzymes (A and B) are those catalyzing the rate limiting step and the reaction necessary for the purine nucleotide cycle,the rate limiting step and the reaction necessary for the purine nucleotide cycle, respectively.respectively. Abbreviations:Abbreviations: PRP: 5-phosphoribosylamine;PRP: 5-phosphoribosylamine; GAR: 5-phosphoribosylglycinamide;GAR: 5-phosphoribosylglycinamide; FGAR: 5-phosphoribosyl-N-formylglycinamideFGAR: 5-phosphoribosyl-N-formylglycinamide FGAM: 5-phosphoribosyl-N-formylglycinamidineFGAM: 5-phosphoribosyl-N-formylglycinamidine AIR: 5-phosphoribosylaminoimidazoleAIR: 5-phosphoribosylaminoimidazole CAIR: 1-(5-phosphoribosyl)-5-amino-4-carboxyimidazoleCAIR: 1-(5-phosphoribosyl)-5-amino-4-carboxyimidazole SAICAR: 1-(5-phosphoribosyl)-4-(N-succinocarboxamide)-5-aminoimidazoleSAICAR: 1-(5-phosphoribosyl)-4-(N-succinocarboxamide)-5-aminoimidazole AICAR: 1-(5-phosphoribosyl)-5-amino-4-imidazolecarboxamideAICAR: 1-(5-phosphoribosyl)-5-amino-4-imidazolecarboxamide FAICAR: 1-(5-phosphoribosyl)-5-formamido-4-imidazolecarboxamideFAICAR: 1-(5-phosphoribosyl)-5-formamido-4-imidazolecarboxamide
  8. 8. The purine base (hypoxanthine) is built upon the ribose by severalThe purine base (hypoxanthine) is built upon the ribose by several amidotransferase and transformylation reactions.amidotransferase and transformylation reactions. The synthesis of IMP requires five moles of ATP, two moles ofThe synthesis of IMP requires five moles of ATP, two moles of glutamine, one mole of glycine, one mole of CO2, one mole ofglutamine, one mole of glycine, one mole of CO2, one mole of aspartate and two moles of formate.aspartate and two moles of formate. IMP represents a branch point for purine biosynthesis, because it canIMP represents a branch point for purine biosynthesis, because it can be converted into either AMP or GMP through two distinct reactionbe converted into either AMP or GMP through two distinct reaction pathways. The pathway leading to AMP requires energy in the form ofpathways. The pathway leading to AMP requires energy in the form of GTP; that leading to GMP requires energy in the form of ATP. TheGTP; that leading to GMP requires energy in the form of ATP. The utilization of GTP in the pathway to AMP synthesis allows the cell toutilization of GTP in the pathway to AMP synthesis allows the cell to control the proportions of AMP and GMP to near equivalence. Thecontrol the proportions of AMP and GMP to near equivalence. The accumulation of excess GTP will lead to accelerated AMP synthesisaccumulation of excess GTP will lead to accelerated AMP synthesis from IMP instead, at the expense of GMP synthesis. Conversely, sincefrom IMP instead, at the expense of GMP synthesis. Conversely, since the conversion of IMP to GMP requires ATP, the accumulation ofthe conversion of IMP to GMP requires ATP, the accumulation of excess ATP leads to accelerated synthesis of GMP over that of AMP.excess ATP leads to accelerated synthesis of GMP over that of AMP.
  9. 9. Regulation of Purine Nucleotide SynthesisRegulation of Purine Nucleotide Synthesis The essential rate limiting steps in purine biosynthesis occur at the firstThe essential rate limiting steps in purine biosynthesis occur at the first two steps of the pathway.two steps of the pathway. The purine nucleotide synthesis is well coordinated to meet theThe purine nucleotide synthesis is well coordinated to meet the cellular demands. The intracellular concentration of PRPP regulatescellular demands. The intracellular concentration of PRPP regulates purine synthesis to a large extent. This in turn is dependent on thepurine synthesis to a large extent. This in turn is dependent on the availability of ribose -5- phosphate and the enzyme PRPP synthetase.availability of ribose -5- phosphate and the enzyme PRPP synthetase. PRPP glutamyl amidotransferase is controlled by feedback mechanismPRPP glutamyl amidotransferase is controlled by feedback mechanism by purine nucleotides. That is, if AMP and GMP are available inby purine nucleotides. That is, if AMP and GMP are available in adequate amounts to meet the cellular needs, their synthesis is turnedadequate amounts to meet the cellular needs, their synthesis is turned off at the amidotransferase reaction.off at the amidotransferase reaction. Another important stage of regulation is in the conversion of IMP toAnother important stage of regulation is in the conversion of IMP to AMP and GMP.AMP inhibits adenylosuccinate synthetase while GMPAMP and GMP.AMP inhibits adenylosuccinate synthetase while GMP inhibits IMP dehydrogenase. Thus, AMP and GMP control theirinhibits IMP dehydrogenase. Thus, AMP and GMP control their respective synthesis from IMP by a feedback mechanism.respective synthesis from IMP by a feedback mechanism.
  10. 10. Catabolism and Salvage of Purine NucleotidesCatabolism and Salvage of Purine Nucleotides Catabolism of the purine nucleotides leadsCatabolism of the purine nucleotides leads ultimately to the production of uric acidultimately to the production of uric acid which is insoluble and is excreted in thewhich is insoluble and is excreted in the urine as sodium urate crystals.urine as sodium urate crystals.
  11. 11. Salvage pathways :Salvage pathways : The synthesis of nucleotides from the purine bases andThe synthesis of nucleotides from the purine bases and purine nucleosides takes place in a series of steps knownpurine nucleosides takes place in a series of steps known as the salvage pathways. The free purine bases---adenine,as the salvage pathways. The free purine bases---adenine, guanine, and hypoxanthine---can be reconverted to theirguanine, and hypoxanthine---can be reconverted to their corresponding nucleotides by phosphoribosylation. Twocorresponding nucleotides by phosphoribosylation. Two key transferase enzymes are involved in the salvage ofkey transferase enzymes are involved in the salvage of purines: adenosine phosphoribosyltransferase (APRT),purines: adenosine phosphoribosyltransferase (APRT), which catalyzes the following reaction:which catalyzes the following reaction: adenine + PRPP <-----> AMP + PPiadenine + PRPP <-----> AMP + PPi and hypoxanthine-guanine phosphoribosyltransferaseand hypoxanthine-guanine phosphoribosyltransferase (HGPRT), which catalyzes the following reactions:(HGPRT), which catalyzes the following reactions: hypoxanthine + PRPP <------> IMP + PPihypoxanthine + PRPP <------> IMP + PPi guanine + PRPP <--------> GMP + PPiguanine + PRPP <--------> GMP + PPi
  12. 12. Clinical Significances of Purine MetabolismClinical Significances of Purine Metabolism Clinical problems associated with nucleotide metabolism in humans areClinical problems associated with nucleotide metabolism in humans are predominantly the result of abnormal catabolism of the purines.predominantly the result of abnormal catabolism of the purines. Clinical manifestations of abnormal purine catabolism arise from theClinical manifestations of abnormal purine catabolism arise from the insolubility of the degradation byproduct, uric acid. Excess accumulation ofinsolubility of the degradation byproduct, uric acid. Excess accumulation of uric acid leads to hyperuricemia, more commonly known as gout. Thisuric acid leads to hyperuricemia, more commonly known as gout. This condition results from the precipitation of sodium urate crystals in the synovialcondition results from the precipitation of sodium urate crystals in the synovial fluid of the joints, leading to severe inflammation and arthritis. Sodium uratefluid of the joints, leading to severe inflammation and arthritis. Sodium urate and uric acid may also precipitate in kidneys and urethras that results in renaland uric acid may also precipitate in kidneys and urethras that results in renal damage and stone formation. Historically , gout was found to be oftendamage and stone formation. Historically , gout was found to be often associated with high living overeating and alcohol consumption. In theassociated with high living overeating and alcohol consumption. In the previous centuries , alcohol was contaminated with lead during itsprevious centuries , alcohol was contaminated with lead during its manufacture and storage. Lead poisoning leads to kidney damage andmanufacture and storage. Lead poisoning leads to kidney damage and decreased uric acid excretion causing gout.decreased uric acid excretion causing gout. The prevalence of gout is about 3 per 1000 persons, mostly affecting males .The prevalence of gout is about 3 per 1000 persons, mostly affecting males . Post-menopausal women are susceptible as men for this disease. Gout is ofPost-menopausal women are susceptible as men for this disease. Gout is of two types- primary and secondary.two types- primary and secondary. Primary goutPrimary gout : It is an inborn error of metabolism due to overproduction of uric: It is an inborn error of metabolism due to overproduction of uric acid. This is mostly related to increased synthesis of purine nucleotides. Theacid. This is mostly related to increased synthesis of purine nucleotides. The following metabolic defects ( enzymes ) associated with primary goutfollowing metabolic defects ( enzymes ) associated with primary gout 1. PRPP synthetase1. PRPP synthetase 2. PRPP glutamylamidotransferase.2. PRPP glutamylamidotransferase. 3. HGPRT deficiency.3. HGPRT deficiency. 4. Glucose -6- phosphatase deficiency4. Glucose -6- phosphatase deficiency 5. Elevation of glutathione reductase5. Elevation of glutathione reductase
  13. 13. 2.2. Secondary goutSecondary gout : Secondary hyperuricemia is due to various disease: Secondary hyperuricemia is due to various disease causing increased synthesis or decreased excretion of uric acid .causing increased synthesis or decreased excretion of uric acid . Increased degradation of nucleic acid ( More uric acid formation ) isIncreased degradation of nucleic acid ( More uric acid formation ) is observed in different cancers (leukemia, lymphomas, polycythemiaobserved in different cancers (leukemia, lymphomas, polycythemia etc ) psoriasis and increased tissue breakdown ( trauma, starvation etcetc ) psoriasis and increased tissue breakdown ( trauma, starvation etc ). The disorders associated with impairment in renal function cause). The disorders associated with impairment in renal function cause accumulation of uric acid which may lead to gout.accumulation of uric acid which may lead to gout. Most forms of gout are the result of excess purine or of a partialMost forms of gout are the result of excess purine or of a partial deficiency in the salvage enzyme, HGPRT. Most forms of gout can bedeficiency in the salvage enzyme, HGPRT. Most forms of gout can be treated by administering the antimetabolite, allopurinol. Thistreated by administering the antimetabolite, allopurinol. This compound is a structural analog of hypoxanthine that strongly inhibitscompound is a structural analog of hypoxanthine that strongly inhibits xanthine oxidase.xanthine oxidase.  Two severe disorders,are associated with defects in purineTwo severe disorders,are associated with defects in purine metabolism: Lesch-Nyhan syndrome and severe combinedmetabolism: Lesch-Nyhan syndrome and severe combined immunodeficiency disease (SCID).immunodeficiency disease (SCID).
  14. 14. GOUTGOUT Once fashionable to associate gout with intelligenceOnce fashionable to associate gout with intelligence people with gout:people with gout:  Isaac NewtonIsaac Newton  Benjamin FrankinBenjamin Frankin  Martin LutherMartin Luther  Charles DarwinCharles Darwin  Samuel JohnsonSamuel Johnson
  15. 15. Non-pharmacological approachesNon-pharmacological approaches Avoid purine rich foods:Avoid purine rich foods:  red meat and organ meat (liver, kidneys)red meat and organ meat (liver, kidneys)  shellfish, anchovies, mackerel, herringshellfish, anchovies, mackerel, herring  meat extracts and graviesmeat extracts and gravies  peas and beans, asparagus, lentilspeas and beans, asparagus, lentils  Weight lossWeight loss
  16. 16. ALLOPURINOL IS A XANTHINE OXIDASE INHIBITORALLOPURINOL IS A XANTHINE OXIDASE INHIBITOR A SUBSTRATE ANALOG IS CONVERTED TO AN INHIBITOR, IN THISA SUBSTRATE ANALOG IS CONVERTED TO AN INHIBITOR, IN THIS CASE “SUICIDE-INHIBITORCASE “SUICIDE-INHIBITOR The drug of choice for the treatment of primary gout is allopurinol. This is a structural analog of hypoxanthine that inhibits the enzyme xanthine oxidase. Allopurinol is oxidized to alloxanthine by xanthine oxidase. Alloxanthine , in turn is a more effective inhibitor of xanthine oxidase This type of inhibition is referred to as suicide inhibition. This inhibition leads to accumulation of hypoxanthine and xanthine. These two compounds are more soluble than uric acid , hence easily excreted. Consumption of plenty of water will also be useful.
  17. 17. Lesch-Nyhan syndrome  This disorder is due to the deficiency of hypoxanthine- guanineThis disorder is due to the deficiency of hypoxanthine- guanine phosphoribosyl transferase ( HGPRT) , an enzyme of purine salvage pathway.phosphoribosyl transferase ( HGPRT) , an enzyme of purine salvage pathway. Lesch-Nyhan syndrome is a sex linked metabolic disorder since the structuralLesch-Nyhan syndrome is a sex linked metabolic disorder since the structural gene for HGPRT is located on the X- chromosome. It affects only the malesgene for HGPRT is located on the X- chromosome. It affects only the males and is characterized by excessive uric acid production (often gouty arthritis )and is characterized by excessive uric acid production (often gouty arthritis ) and neurological abnormalities such as mental retardation , aggressiveand neurological abnormalities such as mental retardation , aggressive behavior , learning disability etc. The patients of this disorder have anbehavior , learning disability etc. The patients of this disorder have an irresistible urge to bite their fingers and lips, often causing self mutilation.irresistible urge to bite their fingers and lips, often causing self mutilation. HGPRT deficiency results in the accumulation of PRPP and decrease in GMPHGPRT deficiency results in the accumulation of PRPP and decrease in GMP and IMP , ultimately leading to increased synthesis and degradation ofand IMP , ultimately leading to increased synthesis and degradation of purines . Neurological symptoms may be related to the dependence of brainpurines . Neurological symptoms may be related to the dependence of brain on the salvage pathway for de novo synthesis of purine nucleotides. Uric acidon the salvage pathway for de novo synthesis of purine nucleotides. Uric acid is not toxic to the brain, since patients with severe hyperuricemia ( notis not toxic to the brain, since patients with severe hyperuricemia ( not related to HGPRT deficiency ) do not exhibit any neurological symptoms.related to HGPRT deficiency ) do not exhibit any neurological symptoms. Allopurinol treatment that helps to decrease uric acid production , has noAllopurinol treatment that helps to decrease uric acid production , has no affect on the neurological manifestations in these patientsaffect on the neurological manifestations in these patients.. Death usuallyDeath usually occurs before patients reach their 20th year.occurs before patients reach their 20th year.
  18. 18. Severe combined immuno deficiency disease :Severe combined immuno deficiency disease :  SCID is caused by a deficiency in the enzyme adenosine deaminase (ADA).SCID is caused by a deficiency in the enzyme adenosine deaminase (ADA). This is the compound responsible for converting adenosine to inosine in theThis is the compound responsible for converting adenosine to inosine in the catabolism of the purines. This deficiency selectively leads to a destruction ofcatabolism of the purines. This deficiency selectively leads to a destruction of B and T lymphocytes, the cells that mount immune responses. In the absenceB and T lymphocytes, the cells that mount immune responses. In the absence of ADA, deoxyadenosine is phosphorylated to yield levels of dATP that are 50-of ADA, deoxyadenosine is phosphorylated to yield levels of dATP that are 50- fold higher than normal. The levels are especially high in lymphocytes, whichfold higher than normal. The levels are especially high in lymphocytes, which have abundant amounts of the salvage enzymes, including nucleoside kinases.have abundant amounts of the salvage enzymes, including nucleoside kinases. High concentrations of dATP inhibit ribonucleotide reductase therebyHigh concentrations of dATP inhibit ribonucleotide reductase thereby preventing other dNTPs from being produced. The net effect is to inhibit DNApreventing other dNTPs from being produced. The net effect is to inhibit DNA synthesis. Since lymphocytes must be able to proliferate dramatically insynthesis. Since lymphocytes must be able to proliferate dramatically in response to antigenic challenge, the inability to synthesize DNA seriouslyresponse to antigenic challenge, the inability to synthesize DNA seriously impairs the immune responses, and the disease is usually fatal in infancyimpairs the immune responses, and the disease is usually fatal in infancy unless special protective measures are taken. A less severe immunodeficiencyunless special protective measures are taken. A less severe immunodeficiency results when there is a lack of purine nucleoside phosphorylase (PNP), anotherresults when there is a lack of purine nucleoside phosphorylase (PNP), another purine- degradative enzyme.purine- degradative enzyme. The deficiency of purine nucleotide phosphorylaseThe deficiency of purine nucleotide phosphorylase is associated with impairment of T- cell function but has no effect on the B- cellis associated with impairment of T- cell function but has no effect on the B- cell functionfunction ..  One of the many glycogen storage diseases von Gierke's disease also leads toOne of the many glycogen storage diseases von Gierke's disease also leads to excessive uric acid production. This disorder results from a deficiency inexcessive uric acid production. This disorder results from a deficiency in glucose 6-phosphatase activity. The increased availability of glucose-6-glucose 6-phosphatase activity. The increased availability of glucose-6- phosphate increases the rate of flux through the pentose phosphate pathway,phosphate increases the rate of flux through the pentose phosphate pathway, yielding an elevation in the level of ribose-5-phosphate and consequentlyyielding an elevation in the level of ribose-5-phosphate and consequently PRPP. The increases in PRPP then result in excess purine biosynthesis.PRPP. The increases in PRPP then result in excess purine biosynthesis.
  19. 19. Pyrimidine Nucleotide BiosynthesisPyrimidine Nucleotide Biosynthesis  Synthesis of the pyrimidines is less complex than that of the purines.Synthesis of the pyrimidines is less complex than that of the purines. The first completed base is derived from 1 mole of glutamine, one moleThe first completed base is derived from 1 mole of glutamine, one mole of ATP and one mole of CO2 (which form carbamoyl phosphate) andof ATP and one mole of CO2 (which form carbamoyl phosphate) and one mole of aspartate. An additional mole of glutamine and ATP areone mole of aspartate. An additional mole of glutamine and ATP are required in the conversion of UTP to CTP.required in the conversion of UTP to CTP. The carbamoyl phosphate used for pyrimidine nucleotide synthesis isThe carbamoyl phosphate used for pyrimidine nucleotide synthesis is derived from glutamine and bicarbonate, within the cytosol, asderived from glutamine and bicarbonate, within the cytosol, as opposed to the urea cycle carbamoyl phosphate derived fromopposed to the urea cycle carbamoyl phosphate derived from ammonia and bicarbonate in the mitochondrion. The urea cycleammonia and bicarbonate in the mitochondrion. The urea cycle reaction is catalyzed by carbamoyl phosphate synthetase I (CPS-I)reaction is catalyzed by carbamoyl phosphate synthetase I (CPS-I) whereas the pyrimidine nucleotide precursor is synthesized by CPS-II.whereas the pyrimidine nucleotide precursor is synthesized by CPS-II. Carbamoyl phosphate is then condensed with aspartate in a reactionCarbamoyl phosphate is then condensed with aspartate in a reaction catalyzed by the rate limiting enzyme of pyrimidine nucleotidecatalyzed by the rate limiting enzyme of pyrimidine nucleotide biosynthesis, aspartate transcarbamoylase (ATCase).biosynthesis, aspartate transcarbamoylase (ATCase). The synthesis of pyrimidines differs in two significant ways from that ofThe synthesis of pyrimidines differs in two significant ways from that of purines. First, the ring structure is assembled as a free base, not builtpurines. First, the ring structure is assembled as a free base, not built upon PRPP. PRPP is added to the first fully formed pyrimidine baseupon PRPP. PRPP is added to the first fully formed pyrimidine base (orotic acid), forming orotate monophosphate (OMP), which is(orotic acid), forming orotate monophosphate (OMP), which is subsequently decarboxylated to UMP. Second, there is no branch insubsequently decarboxylated to UMP. Second, there is no branch in the pyrimidine synthesis pathwaythe pyrimidine synthesis pathway..
  20. 20. Origin of atoms in pyrimidine ringOrigin of atoms in pyrimidine ring
  21. 21. 1. Carbamoyl synthetase 11 2. Aspartate transcarbamylase 3. Dihydrorotase 4. Dihydroorotate dehydrogenase 5. Orotate phosphoribosyl transferase 6. Orotidine -5”-phosphate decarboxylase
  22. 22. Some chemotherapeutic agents block pyrimidine biosynthesis When DNA synthesis is blocked, cells can not divide. Because of this, several important anticancer drugs that block the synthesis of TMP are widely used as chemotheraputic agents. These include pyrimidine analogs such as florourdine as well as antifolates such as methotrexate. One of the unique steps in DNA biosynthesis is the formation of TMP from dUMP by the enzyme thymidylate synthase. The enzyme substrate complex then forms a complex with THF. Once this complex is formed , the methyl transfer occur. FdUMP is a specific suicide inhibitor of thymidylate synthase. This compound can begin the enzymatic conversion into dTMP by forming the enzyme FdUMP complex which can not accept the donated methyl group from methylene TFH nor can it be broken down to release the active enzyme. Flurouridine is used against gastric and uterine cancer. Methotrexate is folic acid analog that bind 1000 fold more tightly to DHFR than does dihydrofolate. Thus , the effectively block the synthesis of THF which in turn limit the formation of methylene THF . In this manner block the synthesis of dTMP and inhibit the synthesis of purine nucleotide. Folate analogs are non specific chemotherapeutic agents. They poison rapidly dividing cells, including those in hair follicles and gut endothelia, causing the loss of hair and GIT side effects of chemotherapy.
  23. 23. Regulation of Pyrimidine BiosynthesisRegulation of Pyrimidine Biosynthesis The regulation of pyrimidine synthesis occurs mainly at the first stepThe regulation of pyrimidine synthesis occurs mainly at the first step which is catalyzed by aspartate transcarbamoylase, ATCase. Inhibitedwhich is catalyzed by aspartate transcarbamoylase, ATCase. Inhibited by CTP and activated by ATP.by CTP and activated by ATP. ATCase, and therefore the activity of CPS-II, is localized to theATCase, and therefore the activity of CPS-II, is localized to the cytoplasm and prefers glutamine as a substrate.cytoplasm and prefers glutamine as a substrate. The CPS-II is activated by ATP and inhibited by UDP, UTP, dUTP, andThe CPS-II is activated by ATP and inhibited by UDP, UTP, dUTP, and CTP.CTP. As in the regulation of purine synthesis, ATP levels regulate pyrimidineAs in the regulation of purine synthesis, ATP levels regulate pyrimidine biosynthesis at the level of PRPP formation. An increase in the level ofbiosynthesis at the level of PRPP formation. An increase in the level of PRPP results in an activation of pyrimidine synthesis.PRPP results in an activation of pyrimidine synthesis. There is also regulation of OMP decarboxylase: this enzyme isThere is also regulation of OMP decarboxylase: this enzyme is competitively inhibited by UMP and, to a lesser degree, by CMP.competitively inhibited by UMP and, to a lesser degree, by CMP. Finally, CTP synthase is feedback-inhibited by CTP and activated byFinally, CTP synthase is feedback-inhibited by CTP and activated by GTP.GTP.
  24. 24. Catabolism and Salvage of Pyrimidine NucleotidesCatabolism and Salvage of Pyrimidine Nucleotides Catabolism of the pyrimidine nucleotides leads ultimately to Beta-alanine (when CMP andCatabolism of the pyrimidine nucleotides leads ultimately to Beta-alanine (when CMP and UMP are degraded) or beta-aminoisobutyrate (when dTMP is degraded) ; NH3 and CO2UMP are degraded) or beta-aminoisobutyrate (when dTMP is degraded) ; NH3 and CO2 The salvage of pyrimidine bases has less clinical significance than that of the purines,The salvage of pyrimidine bases has less clinical significance than that of the purines, owing to the solubility of the by-products of pyrimidine catabolism. However, theowing to the solubility of the by-products of pyrimidine catabolism. However, the salvage pathway to thymidine nucleotide synthesis is especially important in thesalvage pathway to thymidine nucleotide synthesis is especially important in the preparation for cell division. Uracil can be salvaged to form UMP through the concertedpreparation for cell division. Uracil can be salvaged to form UMP through the concerted action of uridine phosphorylase and uridine kinase:action of uridine phosphorylase and uridine kinase: uracil + ribose-1-phosphate <---------> uridine + Piuracil + ribose-1-phosphate <---------> uridine + Pi uridine + ATP ----------> UMP + ADPuridine + ATP ----------> UMP + ADP Deoxyuridine is also a substrate for uridine phosphorylase. Formation of dTMP, byDeoxyuridine is also a substrate for uridine phosphorylase. Formation of dTMP, by salvage of dTMP requires thymine phosphorylase and the previously encounteredsalvage of dTMP requires thymine phosphorylase and the previously encountered thymidine kinase:thymidine kinase: thymine + deoxyribose-1-phosphate <---------> thymidine + Pithymine + deoxyribose-1-phosphate <---------> thymidine + Pi thymidine + ATP ---------> dTMP + ADPthymidine + ATP ---------> dTMP + ADP The salvage of deoxycytidine is catalyzed by deoxycytidine kinaseThe salvage of deoxycytidine is catalyzed by deoxycytidine kinase deoxycytidine + ATP <--------> dCMP + ADPdeoxycytidine + ATP <--------> dCMP + ADP The major function of the pyrimidine nucleoside kinases is to maintain a cellular balanceThe major function of the pyrimidine nucleoside kinases is to maintain a cellular balance between the level of pyrimidine nucleosides and pyrimidine nucleosidebetween the level of pyrimidine nucleosides and pyrimidine nucleoside monophosphates. However, since the overall cellular and plasma concentrations of themonophosphates. However, since the overall cellular and plasma concentrations of the pyrimidine nucleosides, as well as those of ribose-1-phosphate, are low, the salvage ofpyrimidine nucleosides, as well as those of ribose-1-phosphate, are low, the salvage of pyrimidines by these kinases is relatively inefficient.pyrimidines by these kinases is relatively inefficient.
  25. 25. Clinical Significances of Pyrimidine MetabolismClinical Significances of Pyrimidine Metabolism Because the products of pyrimidine catabolism are soluble, fewBecause the products of pyrimidine catabolism are soluble, few disorders result from excess levels of their synthesis or catabolism.disorders result from excess levels of their synthesis or catabolism. Two inherited disorders affecting pyrimidine biosynthesis are theTwo inherited disorders affecting pyrimidine biosynthesis are the result of deficiencies in the bifunctional enzyme catalyzing the last tworesult of deficiencies in the bifunctional enzyme catalyzing the last two steps of UMP synthesis, orotate phosphoribosyl transferase and OMPsteps of UMP synthesis, orotate phosphoribosyl transferase and OMP decarboxylase. These deficiencies result in orotic aciduria that causesdecarboxylase. These deficiencies result in orotic aciduria that causes retarded growth, and severe anemia . Leukopenia is also common inretarded growth, and severe anemia . Leukopenia is also common in orotic acidurias. The disorders can be treated with uridine and/ororotic acidurias. The disorders can be treated with uridine and/or cytidine, which leads to increased UMP production via the action ofcytidine, which leads to increased UMP production via the action of nucleoside kinases. The UMP then inhibits CPS-II, thus attenuatingnucleoside kinases. The UMP then inhibits CPS-II, thus attenuating orotic acid production .orotic acid production .

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