RETNO SINTOWATI

Purin, Pirimidin, Asam nukleat

I. Pendahuluan II. Basa Purin dan Basa Pirimidin III. Nukleosida IV. Nukleotida V. Asam Nukleat

Kepentingan biomedis

kemampuan nukleotid menyerap sinar UV menjadikan sinar UV sbg unsur mutagen yg potent

senyawa analog purin & pirimidin disintesis secara kimiawi digunakan untuk terapi penyakit kanker

Sel yg akan membelah, selnya mengalami replikasi

Katabolisme purin xantin asam urat

I. PendahuluanNukleoprotein

protein majemuk terdapat dalam inti sel penyusun material inti sel tersusun atas : protein: histon / protamin

gugus non protein semua sel hidup mengandung nukleoprotein

KromatinKonstituen penting dalam sel, tersusun atas

nukleoprotein kelainan pembentukan nukleoprotein

perubahan dlm pertumbuhan /reproduksi sel

sebab kromatin pembelahan sel

Nukleosida Purin

Adenosin Guanosin adenosin deaminase purin nukleosida fosforilase

NH3 Inosin Guanin purin nukleosida

fosforilase Hipoxantin Xantin oksidase

Xantin + O2 asam urat urin Xantin oksidase

Penyelamatan Purin Untuk pembentukan nukleotida bbrp jaringan

manusia tgt pd purin /purin ribonukleosida eksogen Ada 2 jalan penyelamatan :1. Fosforibosilasi purin bebas dgn enzim yang

membutuhkan PRPP sbg donor ribosa fosfat2. Fosforilasi nukleosida purin pd gugus 5’-

hidroksi oleh enzim adenosin kinaseContoh :1. Adenin menjadi AMP dg enzim adenin

fosforibosa transferase2. Hipoxantin dan Guanin menjadi IMP dan GMP

dengan enzim hipoxantin-guanin fosforibosa transferase

Nukleosida pirimidin Sitidin Deoksisitidin deaminasi Uridin deaminasi

Deoksiuridin Deoksitimidin

fosforilse

Urasil dan Timin degradasi

-alanin + NH4+ +CO2 degradasi

As. -amino isobutirat + NH4+ +CO2

urine

Banyak tdp pd penderita tumor yang dikhemoterapi ( DNA degradasi )

II. Basa Purin Dan Basa Pirimidin

Basa PurinBasa heterosiklis , molekul planar , aromatik sebag. besar tdp. dlm sel dlm. bentuk asam

nukleatdisintesis secara de novo, dg sumber :

N1 dari as. Aspartat, C6 dr CO2 respirasi, N7 dari glisin, C8 dari N5,N10-metenilFH4,

N3 dan N9 dari nitrogen amida der. Glutamin danC2 dari N-formil -tetrahidroksi folat

Diberi nomor mulai atom N1 kearah kebalikan jarum jam

Basa Pirimidinnama lebih panjang dari purin ttp hy

mgd 6 atom yang heterosiklis, sedang purin

nama pendek mgd 9 atom heterosiklis

Sitosin (S): tdp dlm semua asam nukleat,

kecuali DNA virus ttt

Timin (T) : trtm tdp dlm as nukleatyg mgd gula deoksiribosa (DNA);

sedikit tdp dlm tRNA Urasil (U): tdp dalam RNA

Nitrogenous BasesPlanar, aromatic, and heterocyclicDerived from purine or pyrimidineNumbering of bases is “unprimed”

Nucleic Acid BasesPurines Pyrimidines

SugarsPentoses (5-C sugars)Numbering of sugars is “primed”

Sugars D-Ribose and 2’-Deoxyribose

*Lacks a 2’-OH group

NucleosidesResult from linking one of the sugars with a

purine or pyrimidine base through an N-glycosidic linkage

Purines bond to the C1’ carbon of the sugar at their N9 atoms

Pyrimidines bond to the C1’ carbon of the sugar at their N1 atoms

Nucleosides

Phosphate GroupsMono-, di- or triphosphates

Phosphates can be bonded to either C3 or C5 atoms of the sugar

NucleotidesResult from linking one or more

phosphates with a nucleoside onto the 5’ end of the molecule through esterification

III. Nukleosida dan nukleotidaNukleosida:

- tdr dari basa + d-ribosa gula ribosa terikat

pada N9 basa purin dan pada N1 basa

pirimidin (kecuali U pd C5 ikatan C-C) dan semuanya mrpk ikatan -N-glikosidik

- Nukleosida yang dimanfaatkan untuk menghambat pertumbuhan sel:

- 5-fluorourasil - 5’-iodo-2’-deoksiuridin

- 6-tioguanin - 6-merkaptopurin- 6-azauridin - arabinosil sitosin

NukleotidaMrpk unit struktural as nukleat, mrpk komponen seluler yg mudah dpt diketahui krn menyerap kuat sinar UV, der. basa

purin absorbsinya lebih kuat dari der. basa pirimidinTersusun: - basa purin/ basa pirimidin

- gula (ribosa atau 2-deoksiribosa), - asam fosfatPengikatan fosfat: RNA pd at C3’ dari gulanya DNA pd at C5’ dari gulanya

Basa Nukleosida Nukleotida Singkatan RNA DNA

Adenin Adenosin Asam adenilat A dA

Guanin Guanosin Asam guanilat G dG

Sitosin Sitidin Asam Sitidilat C dCUrasil Uridin Asam uridilat U

dUTimin Timidin Asam timidilat T

dT

Lanjutan nukleotida

- bentuk nukleotida lebih mudah larut dalam air dp nukleosida dan basa bebasnya

Fungsi nukleotida 1. Berperan dlm metabolisme energi

Contoh: ATP - bentuk energi kimia yang diperlukan untuk sel, dihasilkan dari fosforilasi oksidatif atau fosforilasi tingkt

substrat - terlibat dalam kontraksi otot - transport aktif - mempertahankan gradien/ion - membantu sbg donor fosfat untuk

sin- tesis nukleosida 5’-trifosfat substrat untuk reaksi yg dikatalisis oleh RNA dan DNA polimerase

Lanjutan fungsi nukleotida

2. Monomer unit asam nukleat: DNA dan RNA3. Mediator fisiologik: nukleosida dan nukleotida

membantu sbg kunci mediator fisiologik proses metabolik:

- adenosin penting dlm kontrol aliran darah koroner - ADP critical dlm agregasi platelet dan koagulasi darah - cAMP & cGMP: bekerja sbg second messenger - GTP: diperlukan untuk capping mRNA4. Precursor function: GTP adalah prekursor pemben- tukan kofaktor, tetrahidrobiopterin, untuk reaksi

hidroksilasi dan generasi oksida nitrat5. Komponen koenzim: NAD, NADP, FAD

Lanjutan fungsi nukleotida6. Activated intermediate

- nukleotida membantu sbg kunci activated intermediate, diperlukan untuk berbagai reaksi

- UDP-glukosa-kinase intermidiate sintesis glikogen, glikoprotein dll.

7. Allosteric effectors pd metabolisme nukleotida

Sintesis nukleotida - Di dalam sel mamalia disintesis secara de novo- Tidak semua sel mampu mensintesis nukleotid

purin Contoh; sel-sel darah merah- Pada akhir reaksi menghasilkan ITP

prekursor adenosin 5’-monofosfat (AMP) dan guanosin 5’-mono-

- Fosfat (GMP)

Lanjutan sintesis nukleotida

- IMP AMP energinya dari GTP- IMP GMP energinya dari ATP- Apabila terjadi defek dlm reaksi metabolik

menyebabkan kehilangan pengaturan sintesis nukleotida purin over produksi purin asam urat meningkat

Artritis gout Pembentukan deoksi ribonukleotida

- pada keadaan sel tidak proliferasi rendah

- pada saat replikasi sintesis DNA sintesis deoksiribonukleotida meningkat untuk mensuport sintesis DNA

IV. Asam nukleat (Polinukleotid)dibentuk dari ikatan fosfodiester yg

terikat pada C3’- dan C5’ monomer yg berdekatan

ada mol. yg berakhir 5’ dan 3’masing-masing mol. memp. struktur

primernya, contoh 5’-3’p Gp Gp Ap Tp Cp A

Ada 2 macam asam nukleat besar: DNA & RNA

Nucleotide MetabolismPURINE RIBONUCLEOTIDES: formed de novo

i.e., purines are not initially synthesized as free basesFirst purine derivative formed is Inosine Mono-

phosphate (IMP) The purine base is hypoxanthine AMP and GMP are formed from IMP

Purine NucleotidesGet broken down into Uric Acid (a purine)

Buchanan (mid 1900s) showed where purine ring components came from:

N1: Aspartate AmineC2, C8: FormateN3, N9: GlutamineC4, C5, N7: GlycineC6: Bicarbonate Ion

Formyl-THFFormyl-THF

N

NC

CC

CC

N

NH

H

H

H

H

H

Glutamine Glutamine

Aspartate

CO2

Glycine

HN

N

C

C

C

CC

N

NH

H

H

H

H

H

Purine

N

NC

CC

C

H

H

H

H

Pyrimidine

H

H

GlycineGlutamine (2)AspartateN10-Formyl-THF (2)CO2

GlutamineAspartateCO2

(C5H7N4) (C4H6N2)

Purine Nucleotide Synthesis

OH

H

H

CH2

OH OH

H HO

O2-O3P

-D-Ribose-5-Phosphate (R5P)

O

H

H

CH2

OH OH

H HO

O2-O3P

5-Phosphoribosyl--pyrophosphate (PRPP)

P

O

O

O P

O

O

O

ATP

AMP

RibosePhosphatePyrophosphokinase

H

NH2

H

CH2

OH OH

H HO

O2-O3P

-5-Phosphoribosylamine (PRA)

AmidophosphoribosylTransferase

Glutamine + H2O

Glutamate + PPi

H

NH

H

CH2

OH OH

H HO

O2-O3P

CO

H2C NH2

Glycinamide Ribotide (GAR)

GAR Synthetase

Glycine + ATP

ADP+ Pi

H2C

CNH

O

CH

HN

O

Ribose-5-Phosphate

Formylglycinamide ribotide (FGAR)

H2C

CNH

O

CH

HN

HN

Ribose-5-Phosphate

Formylglycinamidine ribotide (FGAM)

THFN10-Formyl-THF

GAR Transformylase

ATP +Glutamine +H2O

ADP +Glutamate + Pi

FGAM Synthetase

HC

CN

CH

N

H2N

Ribose-5-Phosphate

4

5

5-Aminoimidazole Ribotide (AIR)

ATP

ADP + Pi

AIR Synthetase

C

CN

CH

N

H2N

OOC

Ribose-5-Phosphate

4

5

Carboxyamidoimidazole Ribotide (CAIR)

ATP+HCO3

ADP + PiAIR Car boxylase

Aspartate+ ATP

ADP+ Pi

SAICAR Synthetase

AdenylosuccinateLyase

Fumarate

C

CN

CH

N

NH

Ribose-5-Phosphate

4

5

5-Formaminoimidazole-4-carboxamideribotide (FAICAR)

CH2N

O

CH

O

C

CN

CH

N

H2N

Ribose-5-Phosphate

4

5

5-Aminoimidazole-4-carboxamideribotide (AICAR)

CH2N

O

C

CN

CH

N

H2N

CNH

O

HC

COO

CH2

COO

Ribose-5-Phosphate

4

5

5-Aminoimidazole-4-(N-succinylocarboxamide)ribotide (SAICAR)

THF

AICAR Transformylase

N10-Formyl-

THF

Inosine Monophosphate (IMP)

HN

HCN

C

CC

N

CH

N

O

4

5

HH

CH2

OH OH

H HOO2-O3P

IMPCyclohydrolase

H2O

Ribose-P

HCN

C

CC

N

N

CH

N

NH2

Ribose-P

CN

C

CC

HN

N

CH

N

O

HO

Ribose-P

CN

C

CC

HN

N

CH

N

O

H2N

Ribose-P

HCN

C

CC

HN

N

CH

N

O GTPAspartate

GDP + Pi

Fumarate

ATPGlutamine

ADP + Pi

Glutamate

H2O + NAD+

NADH + H+

AMP

GMP

IMP is converted to AMP or GMP

IMP

XMP

Regulatory Control of Purine Nucleotide Biosynthesis

GTP is involved in AMP synthesis and ATP is involved in GMP synthesis (reciprocal control of production)

PRPP is a biosynthetically “central” molecule (why?)ADP/GDP levels – negative feedback on Ribose Phosphate

Pyrophosphokinase Amidophosphoribosyl transferase is activated by PRPP levelsAPRT activity has negative feedback at two sites

ATP, ADP, AMP bound at one site GTP,GDP AND GMP bound at the other site

Rate of AMP production increases with increasing concentrations of GTP; rate of GMP production increases with increasing concentrations of ATP

Regulatory Control of Purine BiosynthesisAt level of IMP production:

Independent controlSynergistic controlFeedforward activation by PRPP

Below level of IMP productionReciprocal control

Total amounts of purine nucleotides controlled

Relative amounts of ATP, GTP controlled

Purine Catabolism and SalvageAll purine degradation leads to uric acid (but it

might not stop there)Ingested nucleic acids are degraded by pancreatic

nucleases, and intestinal phosphodiesterases in the intestine

Group-specific nucleotidases and non-specific phosphatases degrade nucleotides into nucleosidesDirect absorption of nucleosides Further degradation

Nucleoside + H2O base + ribose (nucleosidase) Nucleoside + Pi base + r-1-phosphate (n. phosphorylase)

NOTE: MOST INGESTED NUCLEIC ACIDS ARE DEGRADED AND EXCRETED.

Intracellular Purine CatabolismNucleotides broken into nucleosides by action

of 5’-nucleotidase (hydrolysis reactions)Purine nucleoside phosphorylase (PNP)

Inosine HypoxanthineXanthosine XanthineGuanosine GuanineRibose-1-phosphate splits off

Can be isomerized to ribose-5-phosphate

Adenosine is deaminated to Inosine (ADA : Adeosine deaminase)

Intracellular Purine CatabolismXanthine is the point of convergence for

the metabolism of the purine bases

Xanthine Uric acidXanthine oxidase catalyzes two reactions

Purine ribonucleotide degradation pathway is same for purine deoxyribonucleotides

Adenosine Degradation

Xanthosine Degradation

• Ribose sugar gets recycled (Ribose-1-Phosphate R-5-P ) – can be incorporated into PRPP (efficiency)• Hypoxanthine is converted to Xanthine by Xanthine Oxidase• Guanine is converted to Xanthine by Guanine Deaminase• Xanthine gets converted to Uric Acid by Xanthine Oxidase

GMP AMP ↓ ↓ Guanosin IMP ↓ ↓ ↓ Inosin

↓ Guanin Hipoxantin

↓ ٭↓ Xantin ٭ ↓

Asam urat URIN tmp kerja xantin oxidase, dihambat oleh ٭

Alopurinol

Uric Acid ExcretionHumans – excreted into urine as insoluble crystalsBirds, terrestrial reptiles, some insects – excrete

insoluble crystals in paste form (hewan urikotelik)Excess amino N converted to uric acid

Others – further modification :

Uric Acid Allantoin Allantoic Acid Urea Ammonia

Ekskresi asam urat total man 400-600mg/24jamAspirin dosis tinggi hambat ekskresi & reabsorpsi urat.

Purine Salvage(Jalur penyelamatan purin)Adenine phosphoribosyl transferase (APRT)

Adenine + PRPP AMP + PPi

Hypoxanthine-Guanine phosphoribosyl transferase (HGPRT)

Hypoxanthine + PRPP IMP + PPi

Guanine + PRPP GMP + PPi

(NOTE: THESE ARE ALL REVERSIBLE REACTIONS)

AMP,IMP,GMP do not need to be resynthesized de novo !

Di hati : nukleotida defosforilasi nukleosida

Sering nukleosida diputus basa bebas + gula

nukleosida & basa dibawa ke jar lain refosforilasi mjd nukleotida

Basa bebas diselamatkan dg mereaksikan dg PRPP nukleotida baru

Enzim penghemat basa tsb HGPRTReaksi analog juga terjadi utk pirimidin

A CASE STUDY : GOUTA 45 YEAR OLD MAN AWOKE FROM SLEEP WITH A

PAINFUL AND SWOLLEN RIGHT GREAT TOE. ON THE PREVIOUS NIGHT HE HAD EATEN A MEAL OF FRIED LIVER AND ONIONS, AFTER WHICH HE MET WITH HIS POKER GROUP AND DRANK A NUMBER OF BEERS.

HE SAW HIS DOCTOR THAT MORNING, “GOUTY ARTHRITIS” WAS DIAGNOSED, AND SOME TESTS WERE ORDERED. HIS SERUM URIC ACID LEVEL WAS ELEVATED AT 8.0 mg/dL (NL < 7.0 mg/dL).

THE MAN RECALLED THAT HIS FATHER AND HIS GRANDFATHER, BOTH OF WHOM WERE ALCOHOLICS, OFTEN COMPLAINED OF JOINT PAIN AND SWELLING IN THEIR FEET.

A CASE STUDY : GOUTTHE DOCTOR RECOMMENDED THAT THE MAN

USE NSAIDS FOR PAIN AND SWELLING, INCREASE HIS FLUID INTAKE (BUT NOT WITH ALCOHOL) AND REST AND ELEVATE HIS FOOT. HE ALSO PRESCRIBED ALLOPURINOL.

A FEW DAYS LATER THE CONDITION HAD RESOLVED AND ALLOPURINOL HAD BEEN STOPPED. A REPEAT URIC ACID LEVEL WAS OBTAINED (7.1 mg/dL). THE DOCTOR GAVE THE MAN SOME ADVICE REGARDING LIFE STYLE CHANGES.

Impaired excretion or overproduction of uric acid

Uric acid crystals precipitate into joints (Gouty Arthritis), kidneys, ureters (stones)

Lead impairs uric acid excretion – lead poisoning from pewter drinking gobletsFall of Roman Empire?

Xanthine oxidase inhibitors inhibit production of uric acid, and treat gout

Allopurinol treatment – hypoxanthine analog that binds to Xanthine Oxidase to decrease uric acid production

ALLOPURINOL IS A XANTHINE OXIDASE INHIBITOR

A SUBSTRATE ANALOG IS CONVERTED TO AN INHIBITOR, IN THIS CASE A “SUICIDE-INHIBITOR”

Pyrimidine Ribonucleotide Synthesis Uridine Monophosphate (UMP) is

synthesized firstCTP is synthesized from UMP

Pyrimidine ring synthesis completed first; then attached to ribose-5-phosphate

N1, C4, C5, C6 : AspartateC2 : HCO3

-

N3 : Glutamine amide Nitrogen

2 ATP + HCO3- + Glutamine + H2O

CO

O PO3-2

NH2

Carbamoyl Phosphate

NH2

CNH

CH

CH2

C

COOO

HO

O

Carbamoyl Aspartate

HN

CNH

CH

CH2

C

COOO

O

Dihydroorotate

HN

CNH

C

CHC

COOO

O

Orotate

HN

CN

C

CHC

COOO

O

HH

CH2

OH OH

H HO

O2-O3P

Orotidine-5'-monophosphate(OMP)

HN

CN

CH

CHC

O

O

HH

CH2

OH OH

H HO

O2-O3P

Uridine Monophosphate(UMP)

2 ADP +Glutamate + Pi

CarbamoylPhosphateSynthetase II

AspartateTranscarbamoylase(ATCase)

Aspartate

Pi

H2O

Dihydroorotase

Quinone

ReducedQuinone

DihydroorotateDehydrogenase

PRPP PPi

Orotate PhosphoribosylTransferase

CO2

OMP Decarboxylase

Pyrimidine Synthesis

UMP UTP and CTPNucleoside monophosphate kinase

catalyzes transfer of Pi to UMP to form UDP; nucleoside diphosphate kinase catalyzes transfer of Pi from ATP to UDP to form UTP

CTP formed from UTP via CTP Synthetase driven by ATP hydrolysis

Glutamine provides amide nitrogen for C4 in animals

Regulatory Control of Pyrimidine SynthesisAnimals – regulation at carbamoyl phosphate

synthetase IIUDP and UTP inhibit enzyme; ATP and PRPP

activate itUMP and CMP competitively inhibit OMP

Decarboxylase

*Purine synthesis inhibited by ADP and GDP at ribose phosphate pyrophosphokinase step, controlling level of PRPP also regulates pyrimidines

Caused by defect in protein chain with enzyme activities of last two steps of pyrimidine synthesis

Increased excretion of orotic acid in urine

Symptoms: retarded growth; severe anemia

Only known inherited defect in this pathway (all others would be lethal to fetus)

Treat with uridine/cytidine HOW DOES URIDINE AND CYTIDINE

ADMINISTRATION WORK TO TREAT OROTIC ACIDURIA?

Degradation of PyrimidinesCMP and UMP degraded to bases similarly to

purines DephosphorylationDeaminationGlycosidic bond cleavage

Uracil reduced in liver, forming -alanine Converted to malonyl-CoA fatty acid

synthesis for energy metabolism

Deoxyribonucleotide FormationPurine/Pyrimidine degradation are the same

for ribonucleotides and deoxyribonucleotidesBiosynthetic pathways are only for

ribonucleotide production Deoxyribonucleotides are synthesized from

corresponding ribonucleotides Pd tingkat difosfat,gugus ribosa direduksi mjd

deoksiribosa o/ enz Ribonukleotida reduktase.ex. CTP defosforilasi CDP reduksi dCDP

dCTP defosforilasi dan deaminasi dUMP

Katabolisme pirimidin menghasilkan metabolit yg dpt larut dalam air, yaitu : CO2, NH3, B-alanin, B-aminoisobutirat

Produk katab purin tidak begitu mudah larut, kelarutan ditingkatkan dg alkalinisasi.