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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. Pendahuluan
Nukleoprotein
protein majemuk
terdapat dalam inti sel penyusun material inti sel tersusun atas :

protein: histon / protamin gugus non protein semua sel hidup mengandung nukleoprotein

Kromatin
Konstituen penting dalam sel, tersusun atas

nukleoprotein kelainan pembentukan nukleoprotein perubahan dlm pertumbuhan /reproduksi sel sebab kromatin pembelahan sel

Nukleoprotein
Enzim proteolitik

- Protein sederhana
trtm: histon & protamin

- Asam nukleat
(RNase / DNase)

polinukleotida
polinukleotidase
(fosfoesterse)

mononukleotida
purin/pirimidin nukleotidase

(fosfatase)

nukleosida (fosforilase)
basa purin/basa pirimidin

asam fosfat

gula (ribosa/deoksi ribosa)

Nukleosida Purin
Adenosin
adenosin deaminase

Guanosin
purin nukleosida fosforilase

NH3

Inosin
purin nukleosida fosforilase

Guanin

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 fosfat 2. Fosforilasi nukleosida purin pd gugus 5-hidroksi oleh enzim adenosin kinase Contoh : 1. Adenin menjadi AMP dg enzim adenin fosforibosa transferase 2. Hipoxantin dan Guanin menjadi IMP dan GMP dengan enzim hipoxantin-guanin fosforibosa transferase

Nukleosida pirimidin
Sitidin
deaminasi

Deoksisitidin
deaminasi

Uridin
fosforilse

Deoksiuridin Deoksitimidin 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 Purin
Basa heterosiklis , molekul planar , aromatik sebag. besar tdp. dlm sel dlm. bentuk asam nukleat disintesis 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 dan C2 dari N-formil -tetrahidroksi folat Diberi nomor mulai atom N1 kearah kebalikan jarum jam

Basa Pirimidin
nama 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 nukleat yg mgd gula deoksiribosa (DNA);
sedikit tdp dlm tRNA Urasil (U): tdp dalam RNA

Nitrogenous Bases
Planar, aromatic, and heterocyclic

Derived from purine or pyrimidine


Numbering of bases is unprimed

Nucleic Acid Bases


Purines Pyrimidines

Sugars
Pentoses (5-C sugars)

Numbering of sugars is primed

Sugars
D-Ribose and 2-Deoxyribose

*Lacks a 2-OH group

Nucleosides
Result 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 Groups
Mono-, di- or triphosphates
Phosphates can be bonded to either C3 or C5 atoms of

the sugar

Nucleotides
Result from linking one or more phosphates with

a nucleoside onto the 5 end of the molecule through esterification

III. Nukleosida dan nukleotida


Nukleosida:

- 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

Nukleotida
Mrpk 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 pirimidin Tersusun: - basa purin/ basa pirimidin
- gula (ribosa atau 2-deoksiribosa), - asam fosfat

Pengikatan fosfat:

RNA pd at C3 dari gulanya DNA pd at C5 dari gulanya

Basa
Adenin

Nukleosida
Adenosin

Nukleotida
Asam adenilat

Singkatan
RNA
A

DNA
dA

Guanin
Sitosin Urasil Timin

Guanosin
Sitidin Uridin Timidin

Asam guanilat
Asam Sitidilat Asam uridilat Asam timidilat

G
C U T

dG
dC dU 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 sintesis nukleosida 5-trifosfat substrat untuk reaksi yg dikatalisis oleh RNA dan DNA polimerase

Lanjutan fungsi nukleotida


2. Monomer unit asam nukleat: DNA dan RNA 3. 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 mRNA 4. Precursor function: GTP adalah prekursor pembentukan kofaktor, tetrahidrobiopterin, untuk reaksi hidroksilasi dan generasi oksida nitrat 5. Komponen koenzim: NAD, NADP, FAD

Lanjutan fungsi nukleotida 6. 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 5-3p Gp Gp Ap Tp Cp A

Ada 2 macam asam nukleat besar: DNA & RNA

Nucleotide Metabolism PURINE RIBONUCLEOTIDES: formed de novo


i.e., purines are not initially synthesized as free bases

First purine derivative formed is Inosine Mono-

phosphate (IMP)

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

Purine Nucleotides
Get broken down into Uric Acid (a purine)

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

N1: Aspartate Amine C2, C8: Formate N3, N9: Glutamine C4, C5, N7: Glycine C6: Bicarbonate Ion

CO2 Glycine Aspartate HN Formyl-THF HC N H Glutamine H C C C N H Glutamine H N C Formyl-THF

H C HN HC N H Purine (C5H7N4) C C

H N CH N H

Glycine Glutamine (2) Aspartate N10-Formyl-THF (2) CO2

H C

HN
HC N H

CH CH

Glutamine Aspartate CO2

Pyrimidine (C4H6N2)

Purine Nucleotide Synthesis


OOC
2-

O COO C
ADP + Pi

O3P O CH2 H H

O H

H OH

C4 C H2N
5

N CH N

Aspartate + ATP

HC CH2 COO

N H

C4 C H2N
5

N CH N

OH OH -D-Ribose-5-Phosphate (R5P)
ATP
Ribose Phosphate Pyrophosphokinase

SAICAR Synthetase

Ribose-5-Phosphate Carboxyamidoimidazole Ribotide (CAIR)


AIR Car boxylase ADP + Pi

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


Fumarate
Adenylosuccinate Lyase

AMP

ATP +HCO3

O C

2-

O3P O CH2 H H OH

O H OH

H O

O P O O

O P O O

HC 4 C H2N
5

N CH N
H2N

C4 C H2N
5

N CH N

Ribose-5-Phosphate
5-Phosphoribosyl--pyrophosphate (PRPP)

Ribose-5-Phosphate 5-Aminoimidazole-4-carboxamide ribotide (AICAR)


N10-FormylTHF

5-Aminoimidazole Ribotide (AIR)


ADP + Pi AIR Synthetase ATP

Glutamine + H2O Amidophosphoribosyl


Transferase

Glutamate + PPi

H N H2C CH O NH
O C H

O C H2N

AICAR Transformylase

THF

2-

O3P O CH2 H H OH

O H

NH2

C4 C NH
5

N CH N

C HN

H OH

-5-Phosphoribosylamine (PRA)
Glycine + ATP
GAR Synthetase

Ribose-5-Phosphate Formylglycinamidine ribotide (FGAM)


FGAM Synthetase ADP + Glutamate + Pi ATP + Glutamine + H2O

Ribose-5-Phosphate 5-Formaminoimidazole-4-carboxamide ribotide (FAICAR)


H2O
IMP Cyclohydrolase

ADP + Pi

H 2C O
2-

NH2

H N H2C C
N10-Formyl-THF THF

O C N
4

CH O NH
2-

HN HC N
O3P O CH2 H H OH O

C NH H H OH
GAR Transformylase

CH N
H H OH

C5

O3P O CH2 H H OH

Ribose-5-Phosphate Formylglycinamide ribotide (FGAR)

Glycinamide Ribotide (GAR)

Inosine Monophosphate (IMP)

IMP is converted to AMP or GMP


O C HN HC N C C N CH N Ribose-P H2O + NAD+ N HC N GTP Aspartate GDP + Pi Fumarate NH2 C C C N CH N Ribose-P

IMP

AMP

NADH + H+ O C HN C HO N C C N CH N Ribose-P H2N O C HN C N C C N CH N Ribose-P

ATP Glutamine

ADP + Pi Glutamate

XMP

GMP

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 levels APRT 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 Biosynthesis


At level of IMP production:
Independent control Synergistic control Feedforward activation by PRPP

Below level of IMP production


Reciprocal control

Total amounts of purine nucleotides controlled Relative amounts of ATP, GTP controlled

Purine Catabolism and Salvage


All 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 nucleosides
Direct 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 Catabolism


Nucleotides broken into nucleosides by action of 5-nucleotidase (hydrolysis reactions)
Purine nucleoside phosphorylase (PNP)
Inosine Hypoxanthine Xanthosine Xanthine Guanosine Guanine Ribose-1-phosphate splits off Can be isomerized to ribose-5-phosphate

Adenosine is deaminated to Inosine (ADA : Adeosine deaminase)

Intracellular Purine Catabolism


Xanthine is the point of convergence for the metabolism of the purine bases
Xanthine Uric acid
Xanthine 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 Guanosin
Guanin
Xantin

AMP IMP Inosin Hipoxantin

Asam urat URIN tmp kerja xantin oxidase, dihambat oleh Alopurinol

Uric Acid Excretion


Humans excreted into urine as insoluble crystals
Birds, 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/24jam Aspirin 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 HGPRT Reaksi analog juga terjadi utk pirimidin

A CASE STUDY : GOUT


A 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 : GOUT


THE 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.

Gout 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 goblets
Fall 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 first CTP is synthesized from UMP Pyrimidine ring synthesis completed first; then

attached to ribose-5-phosphate

N1, C4, C5, C6 : Aspartate C2 : HCO3N3 : Glutamine amide Nitrogen

Pyrimidine Synthesis
O

2 ATP + HCO3- + Glutamine + H2O


2 ADP + Glutamate + Pi

C HN CH C N COO
O3P O CH2 H H OH OH O H H

Carbamoyl Phosphate Synthetase II

O
C

C HN CH C N H Orotate
Reduced Quinone

O
PRPP PPi
2-

NH2 O C O PO3-2

C O

Orotate Phosphoribosyl Transferase

COO

Orotidine-5'-monophosphate (OMP)

Carbamoyl Phosphate
Aspartate Aspartate Transcarbamoylase (ATCase)
Dihydroorotate Dehydrogenase

OMP Decarboxylase CO2

Pi

Quinone

O HO NH2 C O N H C CH2 CH COO


HN
H2O

O
HN

C CH CH N

C CH2 CH N H Dihydroorotate COO


2-

C O
O3P O CH2 H H OH OH O H H

C
Dihydroorotase

Carbamoyl Aspartate

Uridine Monophosphate (UMP)

UMP UTP and CTP


Nucleoside 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 Synthesis


synthetase II

Animals regulation at carbamoyl phosphate


UDP and UTP inhibit enzyme; ATP and PRPP activate it UMP 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

Orotic Aciduria
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
ADMINISTRATION WORK TO TREAT OROTIC ACIDURIA?

HOW DOES URIDINE AND CYTIDINE

Degradation of Pyrimidines
CMP and UMP degraded to bases similarly to purines Dephosphorylation Deamination Glycosidic bond cleavage Uracil reduced in liver, forming -alanine Converted to malonyl-CoA fatty acid synthesis for energy metabolism

Deoxyribonucleotide Formation
Purine/Pyrimidine degradation are the same for ribonucleotides and deoxyribonucleotides

Biosynthetic 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.