Ch 32 - Nucleotide Biosynthesis

CHAPTER 32: Nucleotide Metabolism

32.1 An Overview of Nucleotide Biosynthesis and Nomenclature 32.2 The Pyrimidine Ring Is Assembled and Then Attached to a Ribose Sugar 32.3 The Purine Ring Is Assembled on Ribose Phosphate 32.4 Ribonucleotides Are Reduced to Deoxyribonucleotides 32.5 Nucleotide Biosynthesis Is Regulated by Feedback Inhibition 32.6 Disruptions in Nucleotide Metabolism Can Cause Pathological Conditions

PRPP

5-phosphoribosyl-1-pyrophosphate

Purines

Adenine and Guanine

Amino acids serve as precursors to a wide variety of _________ in addition to _____________

Amino acids serve as precursors to a wide variety of biochemicals in addition to proteins.

Ribonucleotides Are Reduced to

Deoxyribonucleotides..

•Thymidylate Is Formed by the Methylation of Deoxyuridylate

Deoxyuridine diphosphate (dUDP) is converted into dUMP. Thymidylate synthase adds a methyl group to dUMP to form thymidylate in a reaction that requires N5,N10-methylenetetrahydrofolate.

Disruptions in Nucleotide Metabolism Can Cause ____________Conditions

Disruptions in Nucleotide Metabolism Can Cause Pathological Conditions

Diagram of the Control of Purine Biosynthesis

Figure 32.10 The control of purine biosynthesis. Feedback inhibition controls both the overall rate of purine biosynthesis and the balance between AMP and GMP production.

Diagram of the de novo Pathway for Pyrimidine Nucleotide Synthesis

Figure 32.3 De novo pathway for pyrimidine nucleotide synthesis. The C-2 and N-3 atoms in the pyrimidine ring come from carbamoyl phosphate, whereas the other atoms of the ring come from aspartate.

Diagram of de novo Purine Biosynthesis

Figure 32.5 De novo purine biosynthesis. (1) Glycine is coupled to the amino group of phosphoribosylamine. (2) N10-Formyltetrahydrofolate (THF) transfers a formyl group to the amino group of the glycine residue. (3) The inner amide group is phosphorylated and then converted into an amidine by the addition of ammonia derived from glutamine. (4) An intramolecular coupling reaction forms a five-membered imidazole ring. (5) Bicarbonate adds first to the exocyclic amino group and then to a carbon atom of the imidazole ring. (6) The imidazole carboxylate is phosphorylated, and the phosphoryl group is displaced by the amino group of aspartate. (7) Fumarate leaves followed by (8) the addition of a second formyl group from N10-formyltetrahydrofolate. (9) Cyclization completes the synthesis of inosinate, a purine nucleotide.

Diagram of the Pathway Generating AMP and GMP

Figure 32.6 Generating AMP and GMP. Inosinate is the precursor of AMP and GMP. AMP is formed by the addition of aspartate followed by the release of fumarate. GMP is generated by the addition of water, dehydrogenation by NAD+, and the replacement of the carbonyl oxygen atom by NH2 derived by the hydrolysis of glutamine.

The Formation of Deoxyribonucleotides

Figure 32.8 The formation of deoxyribonucleotides. Ribonucleotide reductase catalyzes the formation of deoxyribonucleotides from ribonucleotides.

Diagram of Anticancer Drug Targets

Figure 32.9 Anticancer drug targets. Thymidylate synthase and dihydrofolate reductase are choice targets in cancer chemotherapy because the generation of large quantities of precursors for DNA synthesis is required for rapidly dividing cancer cells.

Folic Acid Deficiency Promotes Birth Defects Such as Spina Bifida

Folic acid plays a crucial role in development of the fetus. Insufficient folic acid uptake during pregnancy can result in neural-tube defects such as spina bifida. Spina bifida is characterized by incomplete neural tube formation.

salvage pathway

In salvage pathways, preformed bases are recovered and attached to an activated ribose

In the absence of purines, the enzymes of de novo purine synthesis associate with one another to form complexes called _________that actively catalyze purine _________

In the absence of purines, the enzymes of de novo purine synthesis associate with one another to form complexes called purinosomes that actively catalyze purine synthesis

De novo purine synthesis

In the de novo purine synthesis pathway, the purine ring is sequentially constructed from small molecule donors on a ribose 5-phosphate backbone provided by 5-phosphoribosyl-1-pyrophosphate (PRPP) to form the first purine product, inosine monophosphate (IMP)

32.3 The Purine Ring Is Assembled on Ribose Phosphate DID YOU KNOW?

In the ring form of ribose sugars, the β configuration means that the group attached at C-1 is on the same side of the ring as the —CH2OH group. In the α configuration, the group attached to C-1 and —CH2OH are on opposite sides of the ring.

Nucleotide Biosynthesis Is Regulated by ________________

Nucleotide Biosynthesis Is Regulated by Feedback Inhibition

Nucleotides can be synthesized in which using which two pathways

Nucleotides can be synthesized de novo—from simple precursors—or by salvage pathways.

32.1 An Overview of Nucleotide Biosynthesis and Nomenclature

Nucleotides can be synthesized de novo—from simple precursors—or by salvage pathways. In salvage pathways, preformed bases are recovered and attached to an activated ribose A nucleoside is a base attached to a sugar, and a nucleotide is a nucleoside with one or more phosphoryl groups.

Ribonucleotides Are Reduced to _____________________

Ribonucleotides Are Reduced to Deoxyribonucleotides

The Purine Ring Is Assembled on

Ribose Phosphate

The Purine Ring Is Assembled on Ribose ______

The Purine Ring Is Assembled on Ribose Phosphate

The Pyrimidine Ring Is Assembled and Then Attached to a _________________ Sugar

The Pyrimidine Ring Is Assembled and Then Attached to a Ribose Sugar

32.5 Nucleotide Biosynthesis Is Regulated by Feedback Inhibition... The synthesis of purine nucleotides is controlled by feedback inhibition at several sites

The synthesis of purine nucleotides is controlled by feedback inhibition at several sites 1.Glutamine phosphoribosyl amidotransferase catalyzes the committed step in purine synthesis and is inhibited by a number of ribonucleotides, notably AMP and GMP. 2.AMP inhibits the formation of adenylosuccinate, a precursor to AMP. GMP inhibits the formation of xanthylate, a precursor to GMP. 3.GTP stimulates the synthesis of AMP, whereas ATP stimulates the synthesis of GMP.

Purinosomes

actively catalyze purine synthesis

, the key regulatory enzyme in pyrimidine synthesis.

aspartate transcarbamolyase

de novo pathway

assembles nucleotides from amino acids and other small molecules

Pyrimidines

cytosine, thymine, uracil

Activated ribose (PRPP) + Amino acids + ATP +CO2 = _________ pathway

de novo Figure 32.2 Salvage and de novo pathways. In a salvage pathway, a base is reattached to a ribose, activated in the form of 5-phosphoribosyl-1-pyrophosphate (PRPP). In de novo synthesis, the base itself is synthesized from simpler starting materials, including amino acids. ATP hydrolysis is required for de novo synthesis.

•When cells are grown in medium that includes purines, de novo purine synthesis ________________________

does not occur.

•PRPP is synthesized from ribose 5-phosphate and ATP by PRPP synthetase.

done by the PPP

•Amino acids are precursors to ....

immune system signals, hormones, membrane lipid constituents, and electron carriers, as well as nucleotide bases.

Deoxyribonucleotides

monomers of DNA A deoxyribonucleotide is a nucleotide that contains deoxyribose. They are the monomeric units of the informational biopolymer, deoxyribonucleic acid (DNA). Each deoxyribonucleotide comprises three parts: a deoxyribose sugar (monosaccharide), a nitrogenous base, and one phosphoryl group.

a base attached to a sugar is a

nucleoside

nucleoside with one or more phosphoryl groups

nucleotide

activate ribose + base = ________ pathway

salvage Figure 32.2 Salvage and de novo pathways. In a salvage pathway, a base is reattached to a ribose, activated in the form of 5-phosphoribosyl-1-pyrophosphate (PRPP). In de novo synthesis, the base itself is synthesized from simpler starting materials, including amino acids. ATP hydrolysis is required for de novo synthesis.

32.3 The Purine Ring Is Assembled on Ribose Phosphate explaines

•AMP and GMP are formed from IMP (inosinate) •Nine steps are required to form inosine monophosphate (IMP or inosinate) from 5-phosphoribosyl-1-amine. •IMP is converted into adenylate (AMP) in a pathway that requires GTP. •IMP is also metabolized to guanosine monophosphate (GMP or guanylate) in a pathway that requires ATP.

The Loss of Adenosine Deaminase Activity Results in Severe Combined Immunodeficiency

•Adenosine deaminase, an enzyme important in the degradation of AMP, deaminates adenosine to form inosine. •A deficiency in adenosine deaminase activity is associated with some forms of severe combined immunodeficiency disorder (SCID). Patients suffering from SCID have compromised immune systems and are very susceptible to recurring infections that can result in death.

Amino Acids Acting as Precursors

•Amino acids serve as precursors to a wide variety of biochemicals in addition to proteins. •Amino acids are precursors to immune system signals, hormones, membrane lipid constituents, and electron carriers, as well as nucleotide bases.

32.4 Ribonucleotides Are Reduced to Deoxyribonucleotides.... Several Valuable Anticancer Drugs Block the Synthesis of Thymidylate

•Aminopterin and methotrexate prevent thymidylate synthesis by inhibiting dihydrofolate reductase, the enzyme that regenerates a required cofactor for thymidylate synthase. •Aminopterin and methotrexate are competitive inhibitors of the reductase.

32.3 The Purine Ring Is Assembled on Ribose Phosphate..

•Bases can be recycled by salvage pathways •Two salvage enzymes recycle purine bases. •Adenine phosphoribosyltransferase catalyzes the formation of adenylate, whereas hypoxanthine-guanine phosphoribosyltransferase (HGPRT) catalyzes the formation of inosinate and guanylate.

CLINICAL INSIGHT The Synthesis of Deoxyribonucleotides Is Controlled by the Regulation of Ribonucleotide Reductase..

•Because cancer cells perform much DNA synthesis, ribonucleotide reductase is an attractive chemotherapy target. •Once converted to the diphosphate form in vivo, gemcitabine becomes a suicide inhibitor of the reductase. •Clofarabine and cladribine, upon conversion to their triphosphate forms, allosterically inhibit the reductase.

CLINICAL INSIGHT Salvage Pathways Recycle Pyrimidine Bases

•Because viral thymidine kinase differs from mammalian thymidine kinase, drugs that block viral thymidine kinase are therapeutic agents. Acyclovir, which is converted into a suicide inhibitor by viral thymidine kinase, is used to treat herpes simplex infections.

32.2 The Pyrimidine Ring Is Assembled and Then Attached to a Ribose Sugar (2)

•Carbamoyl phosphate reacts with aspartate to form carbamoylaspartate. This reaction is catalyzed aspartate transcarbamolyase, the key regulatory enzyme in pyrimidine synthesis.

once Carbamoyl Aspartate is produced...it is metabolized to

•Carbamoylaspartate is then metabolized to orotate.

32.4 Ribonucleotides Are Reduced to Deoxyribonucleotides

•Deoxyribonucleotides are synthesized from ribonucleoside diphosphates. •The 2'-hydroxyl group of ribose is replaced by a hydrogen atom in a reaction catalyzed by ribonucleotide reductase, which acts on all four ribonucleotides. In the process of catalysis, sulfhydryls in the reductase become oxidized. For catalysis to continue, the reductase itself must be reduced.

32.4 Ribonucleotides Are Reduced to Deoxyribonucleotides...

•Dihydrofolate is formed by the methylation of deoxyuridine diphosphate •Dihydrofolate reductase regenerates tetrahydrofolate from dihydrofolate.

32.4 Ribonucleotides Are Reduced to Deoxyribonucleotides..

•In turn, reduced thioredoxin is regenerated by electron flow from NADPH. This reaction is catalyzed by thioredoxin reductase, a flavoprotein. •Electrons flow from NADPH to bound FAD of the reductase, to the disulfide of oxidized thioredoxin, then to ribonucleotide reductase, and finally to the ribose unit.

Lesch-Nyhan Syndrome Is a Dramatic Consequence of Mutations in a Salvage-Pathway Enzyme

•Lesch-Nyhan syndrome results if hypoxanthine-guanine phosphoribosyltransferase (HGPRT) is missing. HGPRT is an enzyme in the salvage pathway for guanylate and inosinate. •Lesch-Nyhan patients suffer from compulsive self-destructive behavior, mental deficiency, and gout.

•Kinases Convert Nucleoside Monophosphates into Nucleoside Triphosphates

•Nucleoside monophosphates are converted into diphosphates by specific nucleoside monophosphate kinases. •For instance, UMP kinase forms UDP from UMP. •Nucleoside diphosphate kinases, which have broad specificity, catalyze the interconversion of di- and triphosphates. •Phosphatase convert nucleoside monophosphates into nucleoside

32.6 Disruptions in Nucleotide Metabolism Can Cause Pathological Conditions

•Nucleotide degradation begins with the hydrolytic cleavage of the nucleotide, which yields a nucleoside, in a reaction that is catalyzed by nucleotidases. •Free bases are generated from nucleosides by the phosphorolytic cleavage catalyzed by nucleoside phosphorylases. •The final products of the reaction are the base and ribose 1-phosphate or deoxyribose 1-phosphate.

Once Carbamoylaspartate is metabolized to orotate what happens?

•Orotate reacts with activated ribose in the form of 5-phosphoribosyl-1-pyrophosphate (PRPP) to form orotidylate.

•This reaction is catalyzed by glutamine phosphoribosyl amidotransferase.

•Purines are synthesized on a ribose molecule in the de novo pathway. The initial committed step forms 5-phosphoribosyl-1-amine from PRPP and glutamine where the amine is in the β configuration.

32.3 The Purine Ring Is Assembled on Ribose Phosphate

•Purines are synthesized on a ribose molecule in the de novo pathway. The initial committed step forms 5-phosphoribosyl-1-amine from PRPP and glutamine where the amine is in the β configuration. •This reaction is catalyzed by glutamine phosphoribosyl amidotransferase.

32.5 Nucleotide Biosynthesis Is Regulated by Feedback Inhibition Learning objective 6: Explain how nucleotide synthesis is regulated

•Pyrimidine biosynthesis Is regulated by aspartate transcarbamoylase •Aspartate transcarbamoylase (ATCase) regulates the synthesis of pyrimidine nucleotides. •ATCase is inhibited by CTP, an example of feedback inhibition.

•Pyrimidines are synthesized from _____________, __________________, and ________________, with _________________ often serving as the nitrogen donor.

•Pyrimidines are synthesized from bicarbonate, aspartate, and ammonia, with glutamine often serving as the nitrogen donor.

32.2 The Pyrimidine Ring Is Assembled and Then Attached to a Ribose Sugar

•Pyrimidines are synthesized from bicarbonate, aspartate, and ammonia, with glutamine often serving as the nitrogen donor. •The pyrimidine ring is synthesized first and subsequently attached to an activated ribose. •The first step in pyrimidine synthesis is a three-step reaction to form carbamoyl phosphate catalyzed by carbamoyl phosphate synthetase II (CPS II).

CLINICAL INSIGHT The Synthesis of Deoxyribonucleotides Is Controlled by the Regulation of Ribonucleotide Reductase

•Ribonucleotide reductase from E. coli is a dimer. Each subunit has two allosteric sites: one that regulates enzyme activity and one that regulates substrate specificity. •Binding of dATP to the active site inhibits enzyme activity, an effect reversed by ATP. •Binding of ATP or dATP to the specificity site stimulates the reduction of UDP and CDP. •Binding of thymidine triphosphate (TTP) to the specificity site stimulates reduction of GDP while inhibiting all other pyrimidine reductions. •The increase in dGDP stimulates the reduction of ADP to dADP.

CLINICAL INSIGHT Salvage Pathways Recycle Pyrimidine Bases

•Salvage pathways use preformed pyrimidine bases, recovered from nucleic acid breakdown, to synthesize nucleotides. •Thymine, a product of DNA degradation, is salvaged by first being incorporated into a nucleoside by thymidine phosphorylase. •Thymidine kinase then generates the nucleotide.

32.4 Ribonucleotides Are Reduced to Deoxyribonucleotides Several Valuable Anticancer Drugs Block the Synthesis of Thymidylate

•The folate analog trimethoprim has antibacterial and antiprotozoal activity. •Trimethoprim binds 105-fold less tightly to mammalian dihydrofolate reductase than it does to reductases of susceptible microorganisms.

CLINICAL INSIGHT Several Valuable Anticancer Drugs Block the Synthesis of Thymidylate

•Thymidylate synthase and dihydrofolate reductase are targets of chemotherapy for cancer treatment. •Fluorouracil, an anticancer drug, is converted into fluorodeoxyuridylate (F-dUMP) by cancer cells. F-dUMP is a suicide inhibitor of thymidylate synthase.

•Uridine monophosphate (UMP or uridylate) is generated by the decarboxylation of orotidylate by orotidylate decarboxylase.

•UMP is one of the two major pyrimidines found in RNA. CTP is generated from UMP by the reaction sequence: UMP à UDP à UTP à CTP CTP is Formed by the Amination of UTP

32.3 The Purine Ring Is Assembled on Ribose Phosphate Enzymes of the Purine-Synthesis Pathway Are Associated with One Another in Vivo

•When cells are grown in medium that includes purines, de novo purine synthesis does not occur. •In the absence of purines, the enzymes of de novo purine synthesis associate with one another to form complexes called purinosomes that actively catalyze purine synthesis.

Gout Is Induced by High Serum Levels of Urate

•Xanthine oxidase catalyzes the conversion of xanthine into uric acid, the final common pathway for the degradation of purine nucleotides. •Uric acid ionizes to form urate. •High blood levels of urate induce gout, a painful disease that results from the accumulation of urate crystal in the joints. •Administration of allopurinol, a suicide inhibitor of the oxidase, relieves the symptoms of gout. Purines are then excreted as xanthine and hypoxanthine. •Urate is a potent antioxidant, and urate in the blood may prevent oxidative damage.