NUCLEIC ACIDS &

METABOLISM

Okunowo O Wahab, Ph.D.Biochemistry DepartmentCollege of MedicineUniversity of LagosE-mail: modelprof@yahoo.com

Course Outline Nucleotide & Nucleic Acid Nomenclature

Importance of Nucleotides to Life

Purine Synthesis

i. De Novo Purine Nucleotide Synthesis

ii. Salvage Pathway For Purines

Degradation of Purine Nucleotides

Disorders of Purine Catabolism

Biosynthesis of Pyrimidine Nucleotides

Disorder of Pyimidine Metabolism

Degradation of Pyrimidine Nucleotides

Enzymes Involved in the Digestion of Polynucleotides in the G.I.T

Introduction

Biomolecules found in the nucleus and other parts of a cell includes phosphoric acids and are made up of:

Purine or pyrimidine base.

Sugar moiety (ribose or deoxy ribose)

Phosphate group (Mono,di or tri phosphate)

Nucleotide & Nucleic Acid Nomenclature Purine: Adenine &

Guanine

Pyrimidine: Thymine, Cytosine& Uracil

Nucleoside = Base + sugar

Nucleotide = Base + sugar + phosphate

Nucleotide = Nucleoside + phosphate

DNA & RNA contain the same purine bases: A&G

DNA & RNA contain the pyrimidine Cytosine, but differ in the second pyrimidine base

DNA contain thymine (T) whereas RNA contain Uracil (U).

Examples of nucleotides

DNA & RNA contain the same purine bases: Adenine and Guanine.

DNA & RNA contain the pyrimidine Cytosine, but differ in the second pyrimidine base: DNA contain thymine (T) whereas RNA contain Uracil (U).

Importance Storage & transmission of genetic information

from generation to generation

Precursor for DNA, RNA & Protein synthesis

Carries of activated intermediates in the synthesis of carbohydrate (UDP-glucose), lipids & proteins.

For conservation & utilization of energy (ADP-ATP) released during cellular metabolism

As essential part of coenzymes needed in oxido-reduction reactions (FAD, NAD, NADP)

PURINE SYNTHESIS The atoms of the purine

ring are contributed by a number of compounds, including CO2 which is synthesized in humans from various metabolic pathways including TCA cycle.

The amino acids are non-essential (aspartic acid, glycine, and glutamine) and can by synthesized in the body.

The N10-formyl THF can be synthesized in microbes from the precursor molecules shown in subsequent slide.

Synthesis of N10-formyl THF

This is a major inhibitory step in the series of reactions leading to purine synthesis. This is so because Sulphonamide or sulfa drugs (PABA analog) and Methotrexate (folic acid analog) inhibits the enzymes Dihydropteroate synthetase and Dihyrofolate reductase respectively in THF synthesis. Once THF cannot be synthesized by the intestinal microflora, its role in the synthesis of purine cannot be ascertained.

DE NOVO PURINE NUCLEOTIDE SYNTHESIS

PRPP Synthase is inhibited by purine nucleosides but activated by Pi

PRPP generated also participates in the synthesis of pyrimidines and in the salvage reactions of purines and pyrimidines

The next enzyme in the next reaction; glutamine phosphoribosyl amidotransferase is inhibited by AMP, GMP, IMP by a feedback mechanism.

The purine ring is constructed by a series of reactions that add the donated carbons and nitrogens to a preformed ribose 5-phosphate. Ultimately, Inosine monophosphate is formed. This is then converted to AMP & GMP as shown below.

Biosynthesis of AMP and GMP from IMP with feedback inhibition

IMP, AMP, GMP can be converted to their di or triphophate for example

AMP + ATP Adenylate kinase 2ADP

GMP + ATP Guanylate kinase GDP + ADP

SALVAGE PATHWAY FOR PURINES

Purines that result from normal turnover of cellular nucleic acids or that are obtained from the diet and not degraded can be reconverted into nucleoside triphosphates and used by the body.

This is referred to as “Salvage pathway” for purines.

Two enzymes are involved.

Adenosine phosphoribosyl transferase (APRT) and hyoxanthine guanine phosphoribosyl transferase (HGPRT).

Both enzymes utilizes PRPP as the source of the ribose 5-phosphate group. The release of pyrophosphate (PP) makes these reactions irreversible.

A deficiency of HGPRT causes Lesch-Nyhan Syndrome

DEGRADATION OF PURINE NUCLEOTIDES The end product of purine

catabolism in humans is uric acid. In other primates; allantoin, other mammals further degrade the uric acid to urea or even ammonia.

1. Amino group is removed from AMP to produce IMP, or from Adenine to produce Inosine.

2. IMP & GMP are converted into their respective nucleoside forms inosine & guanosine by the action of 51-nucleotidase.

3. Purine nucleoside phosphorylase converts inosine & guanosine into their respective purine bases; hypoxanthine & xanthine

4. Guanine is deaminated to form xanthine

5. Hypoxanthine is oxidized by xanthine oxidase to xanthine, which is further oxidized by xanthine oxidase to uric acid, the final product of human purine degradation. Uric acid is excreted in the urine.

DISORDERS OF PURINE CATABOLISM ADENOSINE DEAMINASE DEFICIENCY:1. Causes severe combined immunodeficiency involving T-cell & B-cell dysfunction.

This is because there is accumulation of deoxyadenosine which increases adenosylhomocysteine, this substance is toxic to immature lymphocytes resulting in immunocompromised immune system.

2. Extremely large buildups of dATP in red cells (this dATP inhibits ribonucleotide reductase, and therefore DNA synthesis).

3. ADA-deficencient children usually die before 2years of age from overwhelming infection.

PURINE NUCLEOTIDE PHOSPHORYLASE DEFECIENCY: 1. Causes impairment of T-cell function but no apparent effect on B-cell function.

2. It is characterised with decreased uric acid formation, combined with increased levels of purine nucleosides & nucleotides.

3. dGTP is the major nucleotide that accumulates in red cells (dGTP is converted to dATP, an inhibitor of ribonucleotide reductase. dGTP also inhibits the reduction of UDP & CDP)

GOUT: 1. Characterised by hyperuricemia, with recurrent attacks of acute arthritic joint

inflamation caused by uric acid crystals deposition.

2. Primary gout (hyperuricemia) is the form of the disease that is attributable to an in-born error of metabolism, such as overproduction of uric acid.

GOUT Contd.: 3. Secondary hyperuricemia may be caused by other diseases, for

example, cancer, chronic renal insufficiency, HGPRT deficiency, e.t.c.

4. Treatment with allopurinol which inhibits xanthine oxidase leading to accumulation of hypoxanthine & xanthine which are more soluble than uric acid.

HYPOURICEMIA1. Hypouricemia and increased excretion of hypoxanthine and xanthine

are associated with xanthine oxidase deficiency due to a genetic defect or to severe liver damage.

2. Patients with a severe enzyme deficiency may exhibit xanthinuria and xanthine lithiasis

LESCH-NYHAN SYNDROME1. Lesch-Nyhan Syndrome is a inherited disorder associated with a

virtually complete deficiency of hypoxanthine-guanine phosphoribosyltransferase, and therefore inability to salvage hypoxanthine or guanine.

2. This results in excessive production of uric acid, plus characteristic neurological features including self mutilation, involuntary movements, and mental retardation.

3. Enzyme deficiency results in increased levels of PRPP and decreased IMP and GMP, causing increased de novo purine synthesis

Biosynthesis of pyrimidine nucleotides

Unlike purine ring where the ring is constructed on preexisting ribose 5-phosphate, the pyrimidine ring is synthesized before being attached to ribose 5-phosphate, which is donated by PRPP.

The sources of the carbon and nitrogen atoms in the pyrimidine ring are glutamine, CO2, and aspartic acid.

In mammalian cells the synthesis of carbamoyl phosphate, CPSII is inhibited by UTP and activated by ATP & PRPP.

Synthesis of CTP is produced by amination of UTP by CTP synthase.

The biosynthetic pathway for pyrimidine nucleotides

DISORDER OF PYRIMIDINE METABOLISM

OROTIC ACIDURIA1. Orotate phosphoribosyl transferase and OMP

decarboxylase are separate domains of a single polypeptide.

2. Low activities of orotidine phosphate decarboxylase and orotate phosphoribosyltrnsferase result in abnormal growth, megaloblastic anemia, and the excretion of large amounts of orotate in the urine.

3. Feeding in a diet rich in uridine results in improvement of the anemia and decrease excretion of orotate.

DEGRADATION OF PYRIMIDINE NUCLEOTIDES

Unlike purine rings, human cells can open and degrade pyrimidine ring to highly soluble structures such as -alanine and -aminoisobutyrate that can serve as precursors of acetyl coA & Succinyl coA respectively.

Also, pyrimidine can be salvaged and converted to nucleotides by the enzyme pyrimidine phosphoribosyltransferase, which like its counterparts in purine salvage reactions, utilizes PRPP as the source of the ribose-P

ENZYMES INVOLVED IN THE DIGESTION OF POLYNUCLEOTIDES IN THE G.I.T

Ribonucleases and deoxyribonucleasessecreted in pancreatic juice, hydrolyse RNA & DNA primarily to oligonucleotide.

This is further broken down by phosphodiesterase, producing mononucleotides.

Nucleotidase removes Phosphate group hydrolytically.

Nucleosidase degrade the nucleosides to free bases; purines & pyrimidines