Nucleotide Metabolism

Purine metabolism (Overview)

1. Nomenclature/nucleotide structure 2. Extracellular Hydrolysis of Ingested Nucleic Acid 3. De novo synthesis pathways 4. Re-utilization pathways 5. Metabolic diseases of purine Metabolism (Gout, Lesch-Nyhan, SCID)

Other bases that are not part of nucleic acids but have important metabolic roles include orotate, dihydroorotate, hypoxanthine and xanthine

Nucleosides are bases linked to a ribose or deoxyribose at the sugar's 1' position

Nucleotides, the building blocks of nucleic acids, are nucleoside phosphates, with the phosphodiester bond at the sugar's 5' position.

Ribonucleotides are synthesized first, then deoxyribonucleotide are synthesized from modified ribonucleotides. Bases are the nitrogenated components of nucleic acids and may be purines (two rings) or pyrimidines (one ring). Adenine and guanine are the purines in nucleic acids Thymine, cytosine and uracil are the pyrimidines in nucleic acids

Roles of Nucleotides in Cellular Metabolism

The energy currency in metabolic transactions.

The essential chemical links in the response of cells to hormones and other extracellular stimulii. The structural components of an array of enzyme Cofactors and metabolic intermediates.

Biological significance of nucleotide metabolism

Nucleotides make up nucleic acids (DNA and RNA) Nucleotide triphosphates are the energy carriers in cells (primarily ATP) Many metabolic pathways are regulated by the level of the individual nucleotides

Adenine nucleotides are components of many of the coenzymes

Example: cAMP regulation of glucose release

Examples: NAD+, NADP+, FAD, FMN, coenzyme A

Medical significance of nucleotide metabolism

Anticancer agents:
Rapidly dividing cells biosynthesize lots of purines and pyrimidines, but other cells reuse them. Cancer cells are rapidly dividing, so inhibitor of nucleotide metabolism kill them

Antiviral agents
Zidovudine (Retrovir) Lamivudine (Epivir) Valacyclovir (Valtrex)

Structures of nucleotide building blocks and nucleotides

6 5 1 N 2 N 3 PURINE 4 N 8 N 9 H 7 3N 2 N 1 4 5 6

PYRIMIDINE

OH OH P O H OH OH O CH 2 H Base O H H OH

OH P O H OH H O CH 2 H Base O H H

RIBONUCLEOTIDE

DEOXYRIBONUCLEOTIDE

Ribonucleotide phosphate = ribonucleoside

The Purine and Pyrimidine bases found in nucleotides can be synthesized de novo, or can be obtained through salvage pathways that allow the reuse of the preformed bases resulting from normal cell turnover or from the diet.

Structures of Common Purine Bases.

H= 6 oxy purine X= 2,6 dioxy purine

A= 6 amino purine G= 2 amino, 6-oxy purine

Hypoxanthine is an intermediate for Adenine and Guanine

(N source)

Aspartate

(N source)

Glutamine

The common mechanistic them for the conversion of A and G is the conversion of a carbonyl oxygen to an amino group

There are two basic mechanisms to generate purines and pyrimidines

1. DE NOVO BIOSYNTHETIC PATHWAYS

(building the bases from simple building blocks)

2. SALVAGE PATHWAYS

(the reutilization of bases from dietary or catabolic sources)

The biosynthesis of purine (A and G) begins with the synthesis of the ribose-phosphate
Step 1:Activation of ribose-5-phosphate
enzyme: ribose phosphate pyrophosphokinase product: 5-phosphoribosyl-a-pyrophosphate (PRPP) PRPP is also a precursor in the biosynthesis of pyrimidine nucleotides and the amino acids histidine and tryptophan

Step 2 The major regulatory step in purine biosynthesis is the conversion of PRPP to 5-Phosphoribosyl-1-amine

*
Glutamine Glutamate

PRPP

PPi
Amidophosphoribosyl transferase

enzyme: amidophosphoribosyl transferase displacement of pyrophosphate group by glutamine amide nitrogen product: b-5-phosphoribosylamine

Steps 1 and 2 are tightly regulated by feedback inhibition

Amidophosphoribosyl transferase is an important regulatory enzyme in purine biosynthesis. It is strongly inhibited by the end products IMP, AMP, and GMP. This type of inhibition is called FEEDBACK INHIBITION.

Several amino acids are utilized in purine biosynthesis

IMP is the precursor for both AMP and GMP, the base is also called hypoxanthine

Step 2: displacement of pyrophosphate group by glutamine amide nitrogen Step 3: b-phosphoribosylamine reacts with ATP and glycine Step 4: formylation of free a-amino group of GAR Step 5: The amide amino group of a second glutamine is transferred to form formylglycinamidine ribotide (FGAM) Step 6: Closing of the imidazole ring or formation of 5aminoimidazole ribotide Step 7: acquisition of C6 as HCO3Step 8: acquisition of N1 acquired from aspartate in an amide condensation reaction Step 9: Elimination of fumarate Step 10: acquisition of C2 Another formylation reaction Step 11: cyclization or ring closure to form IMP water is eliminated

Purine nucleoside diphosphates and triphosphates: - to be incorporated into DNA and RNA, nucleoside monophosphates (NMPs) must be converted into nucleoside triphosphates (NTPs) - nucleoside monophosphate kinases (adenylate & guanylate kinases)
AMP + ATP 2 ADP accomplished by separate enzymes GMP + ATP GDP + ADP

- nucleoside diphosphate kinase

GDP + ATP

GTP + ADP

same enzyme acts on all nucleotide di & triphosphates nucleoside diphosphate k inase is an enzyme which plays a k ey role in the activation of antiviral nucleosides such as Retrovir/AZ

Last step From IMP to GMP and AMP

Conversion of Hypoxanthine to Adenine/Guanine.

(N source) Aspartate

(N source) Glutamine

The common mechanistic theme for the conversion of A and G is the conversion of a carbonyl oxygen to an amino group

The regulation of purine biosynthesis is a classic example of negative feedback

Inhibited by AMP

AMP Ribose 5-phosphate PRPP Phosphoribosyl amine IMP GMP

Inhibited by IMP, AMP, and GMP

Inhibited by GMP

Salvage pathways for the re-utilization of purines; There are 2 salvage enzymes with different specificity; 1. Adenine phosphoribosyl transferase 2. Hypoxanthine-guanine phosphoribosyl transferase
O
O P O CH2 OH OH O PPi O O

Base (ie Adenine)

P O CH2
OH OH

A
O OH

PPi

OH A-PRT

PRPP + Adenine
HG-PRT

Adenylate

PRPP + Guanine

Guanylate

Nucleic acid degradation

Dietary nucleoprotein is degraded by pancreatic enzymes and tissue nucleoprotein by lysosomal enzymes.
The nucleic acids are hydrolyzed randomly by nucleases to yield a mixture of polynucleotides. polynucleotides are further cleaved by phosphodiesterases (exonucleases) to a mixture of the mononucleotides The nucleotides are hydrolyzed by nucleotidases to give the nucleosides and Pi. The nucleosides undergo phosphorolysis with nucleoside phosphorylases to yield the base and ribose 1-P (or deoxyribose 1-P). The purine and pyrimidine bases released are either degraded or salvaged for reincorporation into nucleotides

Purines in humans are degraded to Urate

Important points:
1. Nucleotides are constantly undergoing turnover! 2. There are many enzymes involved; Nucleotidases Nucleoside phosphorylases Deaminases Xanthine oxidases 3. the final common intermediate in humans is Urate, which is excreted. 4. there are several metabolic disorders resulting from defects in purine catabolism.

NH 2 N N N N

HO
2

NH

4 HN

O N N N

Pi

ribose - 1 - P
HN

O N N N H

ribose Adenosine

Adenosine deaminase (ADA)

ribose Inosine

purine nucleoside phosphorylase

Hypoxanthine xanthine oxidase

O HN NH 2 N N N

Pi

ribose - 1 - P
O HN N N N H H 2O

NH 4

O HN N N H N H

ribose Guanosine

purine nucleoside NH 2 phosphorylase

Guanine deaminase

Xanthine Guanine
O

xanthine oxidase
H N O N H N H

Catabolism of Purine Nucleotides

HN

Uric acid
O