NUCLEOTIDE METABOLISMSITI ANNISA DEVI TRUSDA

Nucleotides are essential for all cellsDNA/RNA synthesisprotein

synthesiscells proliferateCarriers of activated intermediates in the

synthesis of carbohydrate, lipids and protein

Structural component of several essential coenzymes (coA,FAD,NAD+,NADP+)

cAMP,cGMP2nd messenger in signal transduction pathway

Important regulatory compound for many of the pathways of intermediary metabolism, inhibiting/activating key enzimes

Nucleotide structureConsist of:

◦Nitrogenous base : purine & pyrimidine◦Pentose monosaccharide◦1/2/3 phosphate groupsDNA and RNA contain the same purine

bases: A & GPirimidine RNA : U & C DNA : T & CT& U differ by only one methyl group

NucleosidesPentose sugar + Nitrogen Base =

NucleosidesSo, nucleotides = Nucleosides +

PhosphateIf the sugar is ribose : ribonucleosidesIf deoxyribose: deoxyribonucleosidesRibonucleosides of A,G,C,U:

Adenosine,Guanosine,Cytidine,UridineWhat are the deoxyribonucleosides for

A,G,C,T?

Nucleotidesmono,di,tri esters of nucleosides1st phosphate group is attached by an

ester linkage to the 5’OH of the pentosenucleoside 5’phosphate/5’-nucleoside

Type of pentose is added as prefix for nucleotide, can be ribose/deoxyribose e.g: 5’-ribonucleotide/5’-deoxyribonucleotide

1 phosphate group + 5’-carbon of the pentosenucleoside monophosphate(NMP) e.g AMP, CMP

2 or 3 phosphate group added to the nucleosidenucleoside di/triphosphate e.g ADP/ATP

The latter connected to the nucleotide by a high-energy bond

Phosphate groups(-) charge DNA/RNA=nucleic acids

So, what is :◦Nucleoside?◦Nucleotide?◦Nucleic acid?

SYNTHESIS OF PURINE NUCLEOTIDESSource of purine ring: aspartic acid,

glycine, glutamine, CO2,N10-formylTHFSynthesis of 5-phosphoribosyl-1-

pyrophosphate (PRPP)an activated pentose for synthesis of

purine/pirimidine & salvage of purine basescatalyzed by PRPP synthetase, from ATP &

ribose 5-phosphatethis enzyme is activated by inorganic

phosphat (Pi), inhibited by purine nucleotidesthe sugar of PRPP is ribose ribonucleotides

as end product of purine synthetis

Purine synthesis is critical to fetal development, therefore defects in enzymes will result in a nonviable fetus.

PRPP synthetase defects are known and have severe consequences (next slide)

PRPP synthetase superactivity has been documented, resulting in increased PRPP, elevated levels of nucleotides, and increased excretion of uric acid.

Phosphoribosyl Pyrophosphate (PRPP) Synthetase Defects

PRPP deficiency results in convulsions, autistic behavior, anemia, and severe mental retardation.

Excessive PRPP activity causes gout (deposition of uric acid crystals), along with various neurological symptoms, such as deafness.

Synthesis of 5’-phosphoribosylamine

Amide group of glutamine replaces the pyrophosphate group at C1 of PRPP

the enzyme, glutamine:phosphoribosyl pyrophosphate amidotransferase is inhibited by the purine 5’-nucleotides AMP,GMP,IMP (end product)

Committed stepRate of reaction also controlled by

intracellular [] of glutamine and PRPP

Synthesis of inosine monophosphate,the “parent” of purine nucleotide

requires 4 ATP2 steps require N10 –

formyltetrahydrofolate

Conversion of IMP to AMP and GMP

2 step energy requiring pathwaysynthesis of AMP requires GTP as

energy sourcesynthesis of GMP requires ATP

Conversion of nucleoside monophosphates to nucleoside di and triphosphate

AMP + ATP ↔ 2 ADP GMP +ATP ↔ GDP + ADPGDP + ATP ↔ GTP + ADP CDP + ATP ↔ CTP + ADP

Purine Synthesis

DAUR dr IMP AMP & GMP

IMP dehidrogenase

XMP aminase

Adenilosuksinat synthetase

Adenilosuksinat lyase

Salvage Pathway of purines

Purines that result from the normal turnover of cellular nucleic acids/diet can be reconverted into nucleoside triphosphatessalvage pathway

2 enzymes: Adenine phosphoribosyltransferase (APRT), and hypoxanthine-guanine phosphoribosyltransferase (HPRT)

Both needs PRPP as the source of the ribose 5-phosphate

Degradation of Purine Nucleotides Purine Nucleotides from ingested

nucleic acids or turnover of cellular nucleic acids is excreted by humans as uric acid.

Humans excrete about 0.6 g uric acid every 24 hours.

Degradation of dietary nucleic acids occurs in the small intestine by pancreatic enzymes

Digestion of dietary nucleic acidsIn the stomach: low pH denatures

DNA&RNAIn small intestine: break down

phosphodiester bond by endonuclease (pancreas) oligonucleotide

By phosphodiesterase(exonuclease non spesific enzyme) mononucleotide

By phosphomonoesterase (nucleotidase) result: nucleoside and orthophosphate.

Nucleosida phosphorylase result: base and ribose-1-phosphate.

The nucleoside then absorbed by intestinal mucosal cells

If the base or nucleoside is unused, it will be reused in salvage pathways, the base will be degraded:

uric acid ureidopropionic (purin) (pyrimidine).

Diseases associated with purine degradation

GoutElevated uric acid levels

in the bloodUric acid crystals will

form in the extremities with a surrounding area of inflammation. This is called a tophus and is often described as an arthritic “great toe”.

Can be caused by a defect in an enzyme of purine metabolism or by reduced secretion of uric acid into the urinary tract.

tophus

Adenosine Deaminase (ADA) and Purine Nucleoside Phosphorylase (PNP) Deficiency.

accumulation of adenosine wich is converted to its ribonucleotide or deoxyribonucleotide form by cellular kinases

As dATP level rise, ribonucleotide reductase is inhibited↓ production of all deoxyribose containing nucleotidescells cannot make DNA and divide.

Most severe form: severe combined immunodeficiency disease (SCID)lack of T and B cells

A deficiency of either ADA or PNP causes a moderate to complete lack of immune function.

Affected children cannot survive outside a sterile environment.

They may also have moderate neurological problems, including partial paralysis of the limbs.

When a compatible donor can be found, bone marrow transplant is an effective treatment.

Lesch-Nyhan SyndromeHypoxanthine Guanine

Phosphoribosyltransferase (HGPRT) deficiency

X-linked genetic conditionSevere neurologic disease, characterized

by self-mutilating behaviors such as lip and finger biting and/or head banging

Up to 20 times the uric acid in the urine than in normal individuals. Uric acid crystals form in the urine.

Untreated condition results in death within the first year due to kidney failure.

Treated with allopurinol, a competitive inhibitor of xanthine oxidase.

SYNTHESIS OF DEOXYRIBONUCLEOTIDES

Deoxyribonucleotides required for DNA synthesis (2’-deoxyribonucleotides)

Enzyme: ribonucleotide reductase Inhibitor : dATPNeeded a coenzyme : thioredoxinThioredoxin is regenerated by

thioredoxin reductase Regulation of ribonucleotide reduction

is controlled by allosteric feedback mechanisms.

PYRIMIDINE SYNTHESIS AND DEGRADATION

Source of pyrimidine ring: glutamine, CO2, aspartic acid

Synthesis of carbamoyl phosphate

from glutamine & CO2, enzyme: carbamoyl phosphate synthetase II (CPS II), inhibited by UTP

activated by ATP and PRPP

Synthesis of orotic acidformation of

carbamoylaspartatedihydroorotateorotic acid (mind the enzymes!!)

Formation of a pyrimidine nucleotide : orotidine 5’-monophosphate (OMP)the parent of pyrimidine mononucleotide

OMPUridine monophosphate (UMP)Synthesis of uridine triphosphate

and cytidine triphosphateCTP is produced by amination of UTPSynthesis of thymidine

monophosphate from dUMP

Orotat fosforibosiltransferase

Orotidilate dekarboksilase

UMP kinase

CTP synthetase

Nukleosida diphosphat kinase

Pyrimidine Synthesis

Production of Uridine 5’-

monophosphate (UMP) from orotate is

catalyzed by the enzyme UMP

synthase

Orotic AciduriaDeficiency in UMP synthetase activityDue to the demand for nucleotides in the

process of red blood cell synthesis, patients develop the condition of megaloblastic anemia, a deficiency of red blood cells.

Pyrimidine synthesis is decreased and excess orotic acid is excreted in the urine (hence the name orotic aciduria)

Degradation of pyrimidine nucleotides

Unlike the purine rings, which are not cleaved in human cells, the pyrimidine ring can be opened and degraded to highly soluble structures, such as β-alanine, and β-aminoisobutyrate, which can serve as precursors of acetyl coA and succinyl coA

SALVAGE OF PYRIMIDINES

Pyrimidine salvage defects have not been clinically documented

Nucleic Acid Metabolism Overview

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