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NUCLEOTIDENUCLEOTIDE
METABOLISMMETABOLISMBy:By:
Rebecca Asis Villanueva M.D.Rebecca Asis Villanueva M.D.
Associate ProfessorAssociate ProfessorDepartment of Biochemistry & NutritionDepartment of Biochemistry & Nutrition
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NUCLEOTIDENUCLEOTIDE
Ribonucleoside and deoxyribonucleosideRibonucleoside and deoxyribonucleosidephosphatephosphate
Essential for all cellsEssential for all cells
Serve as carriers of activatedServe as carriers of activatedintermediates in the synthesis of someintermediates in the synthesis of somecarbohydrates, lipids and proteinscarbohydrates, lipids and proteins
Structural component of:Structural component of:
coenzyme Acoenzyme AFADFAD
NADNAD
NADPNADP
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Important regulatory compounds forImportant regulatory compounds formany pathways of intermediarymany pathways of intermediary
metabolism inhibiting or activatingmetabolism inhibiting or activatingkey enzymes.key enzymes.
³energy currency´ in the cell ³energy currency´ in the cell
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Nucleotide structureNucleotide structure
Composed of nitrogenous base, aComposed of nitrogenous base, apentose monosaccharide, and onepentose monosaccharide, and one
two or three phosphate groups.two or three phosphate groups. Nitrogen containing bases are:Nitrogen containing bases are:
purinepurine
pyrimidinepyrimidine
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Purine StructurePurine Structure
Guanine
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Pyrimidine structurePyrimidine structure
cytosine
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Pyrimidine structurePyrimidine structure
thymine
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Pyrimidine structurePyrimidine structure
Uracil
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DNA and RNA contain:purine bases: adenosine and guanine
pyrimidine base: cytosine
DNA contains thymine as second pyrimidinebase
RNA contains Uracil as second pyrimidine base
Thymine and Uracil differ only by one methylgroup present in thymine but absent on Uracil
i. e. some viral DNA and transfer RNA
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Presence of unusual base in a nucleotidesequence:
-aid in its recognition by specificenzyme
- protect it from being degraded bynucleases
Base modification:
methylationhydroxymethylationglycosylationacetylation
alterations of the atoms in pyrimidinering
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NucleosidesNucleosides
Addition of pentose sugar to a baseAddition of pentose sugar to a baseproduces a nucleoside.produces a nucleoside.
Adenine AdenosineAdenine AdenosineGuanine GuanosineGuanine Guanosine
Cytosine CytidineCytosine Cytidine
Thymine ThymidineThymine ThymidineUracil UridineUracil Uridine
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Sugar:ribose ribonucleoside
2-deoxyribose deoxyribonucleoside
Carbon and nitrogen atoms in the rings of thebase and the sugar are numbered separately
Atoms in the rings of the bases are numbered1-6 in pyrimidine and 1-9 in purines
Carbons in the pentose are numbered 1-5
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NucleotidesNucleotides
Mono, di or triphosphate esters of Mono, di or triphosphate esters of nucleosidesnucleosides
Phosphate group attached by linkagePhosphate group attached by linkageto the 5¶ to the 5¶--OH of the pentose is calledOH of the pentose is called
nucleoside 5¶ nucleoside 5¶--phosphate orphosphate or
5¶nucleotide5¶nucleotide
Type of pentose denoted prefix in theType of pentose denoted prefix in thenames ³5¶ ribonucleotide and 5¶ names ³5¶ ribonucleotide and 5¶ deoxyribonucleotide´ deoxyribonucleotide´
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Phosphate group are responsible for the
negative changes associated with nucleotidesand nucleic acids
If phosphate group is attached to the 5
carbon of the pentose -Nucleosidemonophosphate (NMP)
i.e. AMP and CMP
If second or third phosphate is added to the
nucleoside - nucleoside diphosphate,triphosphate
i.e. ADP and ATP
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Biomedical ImportanceBiomedical Importance
Biosynthesis of purines and pyrimidinesBiosynthesis of purines and pyrimidinesare regulated and coordinated byare regulated and coordinated byfeedback mechanismfeedback mechanism
Production in quantities vary in times,Production in quantities vary in times,which maybe appropriated withwhich maybe appropriated withphysiologic demandsphysiologic demands
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Genetic diseases associated withGenetic diseases associated with
purine metabolismpurine metabolism
GoutGout
LeschLesch--Nyhan syndromeNyhan syndrome
Adenosine deaminase deficiencyAdenosine deaminase deficiency Purine nucleoside phosphorylasePurine nucleoside phosphorylase
deficiencydeficiency
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Diseases associated withDiseases associated with
pyrimidine catabolismpyrimidine catabolism
Few clinically significant disordersFew clinically significant disorders
Orotic acidemiasOrotic acidemias
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Purine and pyrimidine are dietarily Purine and pyrimidine are dietarily
nonessentialnonessential Human tissue synthesize purine andHuman tissue synthesize purine and
pyrimidine from amphibolicpyrimidine from amphibolicintermediatesintermediates
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Ingested nucleic acids andIngested nucleic acids and
nucleotidesnucleotides Dietarily nonessentialDietarily nonessential
Degraded in the intestinal tract toDegraded in the intestinal tract to
monosaccharide which may bemonosaccharide which may beabsorbed or converted to purine andabsorbed or converted to purine andpyrimidine bases.pyrimidine bases.
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Purine basePurine base
Converted to uric acid by oxidationConverted to uric acid by oxidation
May be absorbed and excreted in theMay be absorbed and excreted in the
urineurine
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DE NOVO PURINEDE NOVO PURINENUCLEOTIDENUCLEOTIDE
SYNTHESISSYNTHESIS
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Atoms of the purine ring areAtoms of the purine ring arecontributed by:contributed by:
--amino acidsamino acids--CO2CO2
--derivatives of tetrahydrofolatederivatives of tetrahydrofolate
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Sources of Individual Atoms inSources of Individual Atoms in
Purine RingPurine Ring
NN
NNN 5N 5
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Synthesis of 5Synthesis of 5--phosphoribosylphosphoribosyl--11--
pyrophosphatepyrophosphate Ribose phosphateRibose phosphate
pyrophosphokinase (PRPPpyrophosphokinase (PRPPsynthetase) is activated by Pi andsynthetase) is activated by Pi andinhibited by purine nucleoside di andinhibited by purine nucleoside di andtriphosphates.triphosphates.
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Synthesis of 5¶Synthesis of 5¶--
phosphoribosylaminephosphoribosylamine
The amide of glutamine replaces theThe amide of glutamine replaces thepyrophosphate group attached topyrophosphate group attached tocarbon 1 of PRPP. The enzymecarbon 1 of PRPP. The enzyme
glutamine: phosphoribosylglutamine: phosphoribosylpyrophosphate amidotransferase ispyrophosphate amidotransferase isinhibited by the purine 5¶ inhibited by the purine 5¶--nucleotidesnucleotidesAMP,GMP and IMP, the end productsAMP,GMP and IMP, the end products
of this pathway. This is theof this pathway. This is thecommitted step in purine nucleotidecommitted step in purine nucleotidebiosynthesisbiosynthesis
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Synthesis of inosineSynthesis of inosine
monophosphate, the ³parent´monophosphate, the ³parent´
purine nucleotidepurine nucleotide
-Requires 4 ATP molecules as an
energy source.
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Inhibitors of purine synthesisInhibitors of purine synthesis
specific for inhibiting the growth of specific for inhibiting the growth of rapidly dividing microorganisms( ex.rapidly dividing microorganisms( ex.Sulfonamides )Sulfonamides )
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Structural analog s of folic acid (ex.Structural analog s of folic acid (ex.Methotrexate) are useMethotrexate) are usepharmacologically to control thepharmacologically to control thespread of cancer by interfering thespread of cancer by interfering thesynthesis of nucleotides, andsynthesis of nucleotides, andtherefore DNA and RNAtherefore DNA and RNA
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Conversion of IMP to AMP andConversion of IMP to AMP and
GMPGMP
The conversion of IMP to either AMPThe conversion of IMP to either AMPor GMP utilizes a twoor GMP utilizes a two--step, energystep, energyrequiring pathway. Note that therequiring pathway. Note that thesynthesis of AMP requires GTP as ansynthesis of AMP requires GTP as anenergy source, whereas theenergy source, whereas thesynthesis of GMP requires ATPsynthesis of GMP requires ATP
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the first reaction in each pathway isthe first reaction in each pathway isinhibited by the end product of thatinhibited by the end product of thatpathway. This provides a mechanismpathway. This provides a mechanismfor diverting IMP to the synthesis of for diverting IMP to the synthesis of the species of purine present inthe species of purine present inlesser amounts.lesser amounts.
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Conversion of nucleosideConversion of nucleoside
monophosphates to nucleoside dimonophosphates to nucleoside di
and triphosphatesand triphosphates
Nucleoside diphosphates areNucleoside diphosphates are
synthesized from the correspondingsynthesized from the correspondingnucleoside monophosphates by basenucleoside monophosphates by base--specific nucleoside monophosphatespecific nucleoside monophosphatekinases.kinases.
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ATP is generally the source of theATP is generally the source of thetransferred phosphate because it istransferred phosphate because it ispresent in higher concentrations thanpresent in higher concentrations thanthe other nucleoside triphosphates.the other nucleoside triphosphates.
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For example, adenylate kinase:For example, adenylate kinase:
AMP + ATPAMP + ATP 2 ADP2 ADP
For example, guanylate kinase:For example, guanylate kinase:GMP + ATPGMP + ATP GDP + ADPGDP + ADP
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Adenylate kinase is particularlyAdenylate kinase is particularlyactive in liver and muscle, where theactive in liver and muscle, where theturnover of energy from ATP is high.turnover of energy from ATP is high.Its function is to maintain anIts function is to maintain anequilibrium among AMP,ADP, andequilibrium among AMP,ADP, andATP:ATP:
2ADP2ADP AMP + ATPAMP + ATP
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Nucleoside diphosphates andNucleoside diphosphates andtriphosphates are interconverted bytriphosphates are interconverted bynucleoside diphosphate kinasenucleoside diphosphate kinase-- ananenzyme that, unlike theenzyme that, unlike themonophosphate kinases, had broadmonophosphate kinases, had broadspecificity.specificity.
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For example,For example, GDP + ATPGDP + ATP GTP + ADPGTP + ADP
For example,For example, CDP + ATPCDP + ATP CTP + ADPCTP + ADP
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SALVAGE P ATHWAY FORSALVAGE P ATHWAY FOR
PURINEPURINE
Purines from the normal turnover of Purines from the normal turnover of cellular nucleic acids or obtainedcellular nucleic acids or obtainedfrom diet that are not degraded canfrom diet that are not degraded can
be reconverted into nucleosidebe reconverted into nucleosidetriphosphatetriphosphate
Energetically much less expensiveEnergetically much less expensivethan complete de novo synthesisthan complete de novo synthesis
Deficiency causesDeficiency causes LeschLesch--NyhanNyhansyndromesyndrome
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Enzymes Involved in SalvageEnzymes Involved in Salvage
PathwayPathway
Adenine phosphoribosyl transferaseAdenine phosphoribosyl transferase
HypoxanthineHypoxanthine--guanineguanine
phosphoribosyl transferasephosphoribosyl transferase
Both utilize PRPP as source of riboseBoth utilize PRPP as source of ribose--55--phosphate groupphosphate group
Release of pyrophosphate makes theseRelease of pyrophosphate makes thesereactions irreversiblereactions irreversible
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Salvage PathwaySalvage Pathway
Two key transferase enzymes are involved in the salvage of
purines: adenosine phosphoribosyltransferase (APRT), which
catalyzes the following reaction:
adenine + PRPP <-----> AMP + PPi
and hypoxanthine-guanine phosphoribosyltransferase (HGPRT),
which catalyzes the following reactions:
hypoxanthine + PRPP <------> IMP +
PPi
guanine + PRPP <--------> GMP + PPi
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LeschLesch--Nyhan SyndromeNyhan Syndrome
Complete deficiency of Complete deficiency of hypoxanthinehypoxanthine--guanineguaninephosphoribosyl transferasephosphoribosyl transferase
Inability to salvage hypoxanthine orInability to salvage hypoxanthine orguanineguanine
Excessive production of uric acidExcessive production of uric acid
Increased levels of PRPPIncreased levels of PRPP
Decreased IMP and GMPDecreased IMP and GMP
Increased de novo
purine synthesis
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DEGRADATION OF PURINEDEGRADATION OF PURINE
NUCLEOTIDESNUCLEOTIDES
Degraded by removal or alterationsDegraded by removal or alterationsof portions of the nucleotideof portions of the nucleotide
End product in human:End product in human:U
ric Acid U
ric Acid Other mammals oxidize uric acid to allantoinOther mammals oxidize uric acid to allantoinwhich can further be degraged to urea orwhich can further be degraged to urea orammoniaammonia
F ti f U i A idF ti f U i A id
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Formation of Uric AcidFormation of Uric Acid
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Degradation of Dietar y NucleicDegradation of Dietar y Nucleic
Acids in Small Intestine Acids in Small IntestineDNA / RNA
Low pH denatures DNA & RNA
Denatured nucleic acids
nucleases
Oligonucleotides
phosphodiesterases
Mononucleotides nucleotidases Nucleosides nucleosidases
Purines / Primidines
mouth
stomach
small intestine
from pancreas
from pancreas
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PYRIMIDINE SYNTHESISPYRIMIDINE SYNTHESIS
Pyrimidine ring synthesized beforePyrimidine ring synthesized beforebeing attached to ribosebeing attached to ribose--55--phosphatephosphate
Sources of carbon and nitrogenSources of carbon and nitrogenatoms:atoms:
GlutamineGlutamine
COCO22
Aspartic acidAspartic acid
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Pyrimidine SynthesisPyrimidine Synthesis
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Synthesis of Carbamoyl PhosphateSynthesis of Carbamoyl Phosphate
Committed stepCommitted step
Catalyzed by: Carbamoyl phosphateCatalyzed by: Carbamoyl phosphate
synthetaseII
synthetaseII
Inhibited by UTPInhibited by UTP
Activated by ATP and PRPPActivated by ATP and PRPP
cytosoliccytosolic UsesUses --amide group of amine asamide group of amine as
source of nitrogensource of nitrogen
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Synthesis of Orotic AcidSynthesis of Orotic Acid
Second step: formation of carbamoylSecond step: formation of carbamoylaspartateaspartate
Catalyzed by aspartate transcarbamoylaseCatalyzed by aspartate transcarbamoylase
Pyrimidine ring closed byPyrimidine ring closed bydihydroorotase resulting todihydroorotase resulting todihydroorotate oxidized to oroticdihydroorotate oxidized to orotic
acidacid
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Formation of Pyrimidine NucleotideFormation of Pyrimidine Nucleotide
Complete pyrimidine ring convertedComplete pyrimidine ring convertedto orotidine 5¶ to orotidine 5¶--monophosphatemonophosphate
PRPPPRPP ±± ribose 5ribose 5--phosphate donorphosphate donor
Orotate phosphoribosyl transferaseOrotate phosphoribosyl transferaseproduces OMP with release of produces OMP with release of pyrophosphatepyrophosphate
Biologically irreversibleBiologically irreversible
OMP converted to UMPOMP converted to UMP By Orotidylate decarboxylaseBy Orotidylate decarboxylase
Deficiency: Orotic aciduriaDeficiency: Orotic aciduria
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Synthesis of Uridine TriphosphateSynthesis of Uridine Triphosphate
and Cytidine Triphosphateand Cytidine Triphosphate
Amination of UTPAmination of UTP By CTP synthaseBy CTP synthase
NitrogenNitrogen
provided byprovided byglutamineglutamine
Synthesis of CTP from UTP
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DEGRADATION OF PYRIMIDINEDEGRADATION OF PYRIMIDINE
NUCLEOTIDESNUCLEOTIDES
Pyrimidine rings can be degraded toPyrimidine rings can be degraded tohighly soluble structureshighly soluble structures
Such asSuch as -- alanine & alanine & --aminoisobutyrateaminoisobutyrate
Can be salvaged and converted intoCan be salvaged and converted intonucleotidesnucleotides
By pyrimidine phosphoribosyltransferaseBy pyrimidine phosphoribosyltransferase
Utilizes PRPP as source of riboseUtilizes PRPP as source of ribose--PP
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CONVERSION OF RIBONUCLEOTIDES TOCONVERSION OF RIBONUCLEOTIDES TO
DEOXYRIBONUCLEOTIDESDEOXYRIBONUCLEOTIDES
RIBONUCLEOTIDES
Use as building blocks in RNA synthesis.as nucleotide carriers of other compound
The nucleotides required for DNA synthesis are2· ² deoxyribonucleotides, which are produced fromribonucleoside diphosphate
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A. RIBONUCLEOTIDE REDUCTASE A. RIBONUCLEOTIDE REDUCTASE
Ribonucleotide reductaseRibonucleotide reductase (ribonucleoside(ribonucleosidediphosphate reductase )diphosphate reductase )
A multi subunit enzyme ( two identical B1A multi subunit enzyme ( two identical B1subunits and two identical B2 subunits) that issubunits and two identical B2 subunits) that isspecific for reduction of nucleoside diphosphatesspecific for reduction of nucleoside diphosphates(dADP,dGDP,dCDP, and dUDP )(dADP,dGDP,dCDP, and dUDP )
Immediate donors of hydrogen atom needed forImmediate donors of hydrogen atom needed forthe reduction of the 2¶ the reduction of the 2¶--hydroxyl group are twohydroxyl group are two
sulfhydryl groups on the enzyme itself sulfhydryl groups on the enzyme itself²²whichwhichduring rreaction forms a disulfide bondduring rreaction forms a disulfide bond
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CONVERSION OF RIBONUCOTIDES TOCONVERSION OF RIBONUCOTIDES TO
DEOXYRIBONUCLEOTIDESDEOXYRIBONUCLEOTIDES
O-P-O-P-P-CH2
H
OO O
O O
O
OH OH
H H
H
BASE
RIBONUCLEOSIDEDIPHOSPHATE
DEOXYRIBONUCLEOSIDE
DIPHOSPHATE
O-P-O-P-O-CH2
H
H H
HOH
BASE
H
O
O O
O O
R ibonucleotide diphosphate
Thioredoxin (2 SH)(reduced)
NADP+
Deoxyribonucleotide diphosphate
Thioredoxin ( S ± S)(oxidized)
NADPH + H+
H20
Ribonucleotide reductase
Thioredoxin reductase
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1.Regeneration of reduced enzyme :1.Regeneration of reduced enzyme :
for fibonucleotide reductase to continue to produce deoxyRibonucleotides, the disulfide bong created during production of the2¶-deoxy carbon must be reduced.
The source of reducting equivalents is a peptide co enzyme of Ribonucleotide reductase , THIOREDOXIN.
Thioredoxin ± contains 2 Cysteine residues separated by two aminoacids in peptide chain.
The 2 sulfhydryl groups of thioredoxin donate their hydrogen atoms toribonucleotide reductase in the process of forming a disulfide bond.
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2. Regeneration of reduced Thioredoxin :2. Regeneration of reduced Thioredoxin :
Thioredoxin must be converted back to its reduced form inOrder to continue its function.
The necessary equivalents are provided by NADPH + H, andthe reaction is catalyzed by Thioredoxin reductase.
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B. REGULATION OF DEOXYRIBONUCLEOTIDEB. REGULATION OF DEOXYRIBONUCLEOTIDE
SYNTHESISSYNTHESIS
Ribonucleotide reductase ± responsible for maintaining a balance supplyOf the deoxyribonuclotides required for DNA synthesis.
to achieve this, the regulation of the enzyme is complex. In additionto the single active site, there are two sites on the enzyme involvedIn regulating this activity.
1. Activity Site- the binding of dATP to an Allosteric site (known as the activity site),
inhibits the overall catalytic activity of the enzyme.
2. Substrate specificity site :- the binding of nucleoside triphosphate to an additional allosteric site
(known as the substrate specificity site ) on the enzyme regulatesSubstrate specificity, causing an increase in the conversion of Ribonucleotides to deoxyribonucleotides as they are required for DNAsynthesis.
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REGULATION OF RIBONUCLEOTIDE REDUCTASEREGULATION OF RIBONUCLEOTIDE REDUCTASE
B2subunit
B2subunitR ibonucleoside
Diphosphate(NDP)
DeoxyribonucleosideDiphosphate (dNDP)
B1
subunit
B1
subunit
SUBSTRATE SPECIFICITY SITE
ACTIVITY SITES
SH SH SH SH
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SYNTHESIS OF TH YMIDINESYNTHESIS OF TH YMIDINE
MONOPHOSPHATE FROM dUMPMONOPHOSPHATE FROM dUMP
dUMP is converted to dTMP by thymidylate synthetase, w/cUtilizes N5, N10 ± methylene tetrahydrofolate as the source of themethyl group.
this is unusual reaction in that tetrahydrofolate (THF) contributesnot only a carbon unit but also TWO hydrogen atoms for thepteridine ring, resulting in the oxidation of THF to dihydrofolate
(DHF)
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Inhibitors of Thymidylate synthetase include thymine analogssuch as 5- fluoracil.
DHF can be reduced to THF by dihyrofolate reductse, an enzymeThat is inhibited in the presence of drugs such as methotrexate.
by decreasing the supply of THF, these folate analogs not onlyInhibit purine synthesis, but by preventing methylation of dUMPto dTMP, they also lower the cellular concentration of this essentialComponent of DNA
DNA synthesis is therefore inhibited and cell growth slowed.
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Disorders of PurineDisorders of Purine
Catabolism:Catabolism:I.I. GOUT (Metabolic Disorder of Purine GOUT (Metabolic Disorder of Purine
Catabolism)Catabolism)
Various genetic defects in PRPP synthetaseVarious genetic defects in PRPP synthetasepresent clinically as Gout.present clinically as Gout.
Results in overproduction and overexcretion of Results in overproduction and overexcretion of purine catabolites or resistance to feedbackpurine catabolites or resistance to feedbackinhibition.inhibition.
e.g. An elevated Vmax increased affinity fore.g. An elevated Vmax increased affinity forribose 5ribose 5--phosphate.phosphate.
Gouty ArthritisGouty Arthritis
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Disorders of PurineDisorders of Purine
Catabolism:Catabolism:II.II. LeschLesch-- NyhanNyhan SyndromeSyndrome AnAn overproductionoverproduction hyperuricimiahyperuricimia characterizedcharacterized byby
frequentfrequent episodesepisodes of of uricuric acidacid lithiasislithiasis andand aa bizarrebizarresyndromesyndrome of of self self mutilationmutilation
TheThe accompanyingaccompanying riserise inin intracellularintracellular PRPPPRPP resultsresultsinin purinepurine overproductionoverproduction..
MutationsMutations thatthat decreasedecrease oror abolishabolish hypoxanthinehypoxanthine--
guanineguanine phosphoribosyltransferasephosphoribosyltransferase activityactivity includeincludedeletions,deletions, frameshiftframeshift mutations,mutations, basebase substitutions,substitutions,andand aberrantaberrant mRNAmRNA splicingsplicing..
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Disorders of PurineDisorders of Purine
Catabolism:Catabolism:III.III. VonVon Gierke¶sGierke¶s DiseaseDisease
PurinePurine overproductionoverproduction andand hyperuricemiahyperuricemia(glucose(glucose--66--phosphatasephosphatase deficiency)deficiency) occursoccurs
secondarysecondary toto enhancedenhanced generationgeneration of of thethe PRPPPRPPprecursorprecursor riboseribose 55--phosphatephosphate..
IV.IV. HypouricemiaHypouricemia
HypouricaemiaHypouricaemia andand increasedincreased excretionexcretion of of hypoxanthinehypoxanthine andand xanthinexanthine areare associatedassociatedwithwith xanthinexanthine oxidaseoxidase deficiencydeficiency duedue totogeneticgenetic defectdefect oror toto severesevere liverliver damagedamage..
ff
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Disorders of PurineDisorders of Purine
Catabolism:Catabolism:V.V. AdenosineAdenosine DeaminaseDeaminase & & PurinePurine
NucleosideNucleoside PosphorylasePosphorylase DeficiencyDeficiency AA deficiencydeficiency whichwhich isis associatedassociated withwith anan
immunodeficiencyimmunodeficiency diseasedisease inin whichwhich bothboth thymusthymusderivedderived lymphocyteslymphocytes (T(T cells)cells) andand bonebone marrowmarrow--derivedderived lymphocyteslymphocytes (B(B cells)cells) areare spursespurse andanddysfunctionaldysfunctional..
PurinePurine nucleosidenucleoside phosphorylasephosphorylase deficiencydeficiency isis
associatedassociated withwith aa severesevere deficiencydeficiency of of TT cellscells butbutapparentlyapparently normalnormal BB cellcell functionfunction..
ImmuneImmune dysfunctionsdysfunctions appearappear toto resultresult fromfromaccumulationaccumulation of of dGTPdGTP andand dATP,dATP, whichwhich inhibitinhibitribonucleotideribonucleotide reductasereductase andand therebythereby depletedeplete cellscells of of
DNADNA recursorsrecursors..
Overproduction of Pyrimidine CatabolitesOverproduction of Pyrimidine Catabolites
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Overproduction of Pyrimidine CatabolitesOverproduction of Pyrimidine Catabolites
is only rarely associated with clinicallyis only rarely associated with clinically
significant abnormalitiessignificant abnormalities Since the end products of pyrimidine catabolismSince the end products of pyrimidine catabolism
are highly waterare highly water--soluble, pyrimidinesoluble, pyrimidineoverproduction results in few clinical signs oroverproduction results in few clinical signs or
symptoms.symptoms.
In hyperuricemia associated with severeIn hyperuricemia associated with severeoverproduction of PRPP, there is overproductionoverproduction of PRPP, there is overproduction
of pyrimidine nucleotides and increased excretionof pyrimidine nucleotides and increased excretionof ßof ß--alanine. Sincealanine. Since N5 N5 ,, N10N10 ±±methylenemethylene--tetrahydrofolate is required for thymidylatetetrahydrofolate is required for thymidylatesynthesis, disorders of folate and vitamine B12synthesis, disorders of folate and vitamine B12metabolism results in deficiencies of TMP.metabolism results in deficiencies of TMP.
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Clinically SignificantClinically Significant
AbnormalitiesAbnormalities
I.I. Orotic AciduriasOrotic Acidurias
TheThe orotic aciduriasorotic acidurias that accompaniesthat accompaniesReye¶s syndromeReye¶s syndrome probably is consequence of probably is consequence of the inability of severely damaged mitochondriathe inability of severely damaged mitochondriato utilize carbamoyl phosphate which thento utilize carbamoyl phosphate which thenbecomes available for cytosolic overproductionbecomes available for cytosolic overproductionof orotic acid.of orotic acid.
Type I orotic aciduriaType I orotic aciduria reflects a deficiency of reflects a deficiency of
both orotate phosphoribosyltransferase andboth orotate phosphoribosyltransferase andorotidylate decarboxylase.orotidylate decarboxylase.
The rarerThe rarer Type II orotic aciduriaType II orotic aciduria is due to ais due to adeficiency only of orotidylate decarboxylase.deficiency only of orotidylate decarboxylase.
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Clinically SignificantClinically Significant
AbnormalitiesAbnormalities
II.II. Deficiency of a Urea Cycle Enzyme Deficiency of a Urea Cycle Enzyme results in Excretion of Pyrimidine results in Excretion of Pyrimidine PrecursorsPrecursors
Increased excretion of orotic acid, uracil, andIncreased excretion of orotic acid, uracil, anduridine accompanies a deficiency in liveruridine accompanies a deficiency in livermitochondrialmitochondrial ornithine ornithine transcarbamoylasetranscarbamoylase..
Excess carbamoyl phosphate exits to theExcess carbamoyl phosphate exits to thecytosol, where it stimulates pyrimidinecytosol, where it stimulates pyrimidinenucleotide biosynthesis.nucleotide biosynthesis.
The resultingThe resulting mild orotic aciduriamild orotic aciduria is increasedis increasedby high nitrogen containing food.by high nitrogen containing food.
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Clinically SignificantClinically Significant
AbnormalitiesAbnormalities
III.III. Drugs May Precipitate Orotic Drugs May Precipitate Orotic AciduriaAciduria
Allopurinol Allopurinol ,, an alternative substrate foran alternative substrate fororotate phosphoribosyltransferase, competesorotate phosphoribosyltransferase, competes
with orotic acid.with orotic acid. The resulting nucleotide product also inhibitsThe resulting nucleotide product also inhibits
orotidylate decarboxylase, resulting in oroticorotidylate decarboxylase, resulting in oroticaciduria and ortidinuria.aciduria and ortidinuria.
66--Azauridine Azauridine, following conversion to 6, following conversion to 6--azauridylate, also competitively inhibitsazauridylate, also competitively inhibits
orotidylate decarboxylaseorotidylate decarboxylase, enhancing, enhancing
exretion of orotic acid and orotidineexretion of orotic acid and orotidine
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INHIBITORS OF PURINE METABOLISMINHIBITORS OF PURINE METABOLISM
(CANCER CHEMOTHERAP Y)(CANCER CHEMOTHERAP Y)
III.III. The turnover of nucleic acids in malignant of itsThe turnover of nucleic acids in malignant of itsformation in these cells may offer a therapeutic value.formation in these cells may offer a therapeutic value.Since most if not. All of these ³inhibitor drugs´ are nonSince most if not. All of these ³inhibitor drugs´ are non--specific, then there is a possibility that it can also affectspecific, then there is a possibility that it can also affectnormal tissues which are rapidly undergoing mitoticnormal tissues which are rapidly undergoing mitotic
cycles like the hematopoietic tissues hence should becycles like the hematopoietic tissues hence should beadministered with proper caution.administered with proper caution.
1.1. 66--Mercaptopurine (6MP)Mercaptopurine (6MP)
This is an antitumor drug which is an analougue of adenineThis is an antitumor drug which is an analougue of adeninemetabolized to a ribonucleotide by the APRT salvagemetabolized to a ribonucleotide by the APRT salvagepathway thus inhibiting the conversion of IMP to GMP orpathway thus inhibiting the conversion of IMP to GMP or
AMP. It also inhibits the rate limiting step of the purineAMP. It also inhibits the rate limiting step of the purinede novo pathway. Simultaneous administration withde novo pathway. Simultaneous administration withallopurinol potentiates its effect as 6allopurinol potentiates its effect as 6--MercaptopurineMercaptopurinewill not be degraded and therefore will delay itswill not be degraded and therefore will delay itsinactivation.inactivation.
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INHIBITORS OF PURINE METABOLISMINHIBITORS OF PURINE METABOLISM
(CANCER CHEMOTHERAP Y)(CANCER CHEMOTHERAP Y)
2. Adenosine arabinoside (the sugar is replaced by2. Adenosine arabinoside (the sugar is replaced byan arabinose)an arabinose)
This is used as an antiviral and an antitumor drugThis is used as an antiviral and an antitumor drugin man. It inhibits DNA polymerase after itsin man. It inhibits DNA polymerase after itsconversion to the triphosphate form.conversion to the triphosphate form.
3. Azaserine3. Azaserine
An analogue of glutamine, thus inhibits theAn analogue of glutamine, thus inhibits theincorporation of N3 and N9 into the purineincorporation of N3 and N9 into the purine
ring (de novo biosynthesis), inhibits formationring (de novo biosynthesis), inhibits formationof GMP from IMP, CTP from UTPof GMP from IMP, CTP from UTP--all reactionsall reactionsrequiring the entry of glutamine.requiring the entry of glutamine.
INHIBITORS TO PYRIMIDINE NUCLEOTIDEINHIBITORS TO PYRIMIDINE NUCLEOTIDE
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INHIBITORS TO P YRIMIDINE NUCLEOTIDEINHIBITORS TO P YRIMIDINE NUCLEOTIDE
METABOLISM (CANCER AND VIRALMETABOLISM (CANCER AND VIRAL
CHEMOTHERAP Y)CHEMOTHERAP Y)
1.1. Aminopterine/Amithopterine/MethotrexateAminopterine/Amithopterine/Methotrexate
--Inhibits dihydrofolate reductase.Inhibits dihydrofolate reductase.
2. 5 Flurouracil (5 FU)2. 5 Flurouracil (5 FU)
-- An analogue of thymine used in the treatment of solidAn analogue of thymine used in the treatment of solidtumors. It is converted to the monophosphatetumors. It is converted to the monophosphatenucleotide form via the salvage pathway. Eventually itnucleotide form via the salvage pathway. Eventually itis converted to the deoxynucleotide form and binds tois converted to the deoxynucleotide form and binds tothymidylate synthetase, thereby inhibiting thethymidylate synthetase, thereby inhibiting theformation of TMP. As a deoxytriphosphate form, it canformation of TMP. As a deoxytriphosphate form, it canbe incorporated into RNA and inhibits the formation of be incorporated into RNA and inhibits the formation of mature RNA (a step important in translation)mature RNA (a step important in translation)
3. 53. 5--IodouracilIodouracil
-- Functions as an analogue of thymidine, which whenFunctions as an analogue of thymidine, which whenincorporated into DNA bonds with C rather than A, thusincorporated into DNA bonds with C rather than A, thuscausing misreading of the strand.causing misreading of the strand.
INHIBITORS TO PYRIMIDINE NUCLEOTIDEINHIBITORS TO PYRIMIDINE NUCLEOTIDE
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INHIBITORS TO P YRIMIDINE NUCLEOTIDEINHIBITORS TO P YRIMIDINE NUCLEOTIDE
METABOLISM (CANCER AND VIRALMETABOLISM (CANCER AND VIRAL
CHEMOTHERAP Y)CHEMOTHERAP Y)
4. Cytosine arabinoside (sugar is replaced to an arabinose)4. Cytosine arabinoside (sugar is replaced to an arabinose)
--used in the treatment of leukemias. It ingibits DNAused in the treatment of leukemias. It ingibits DNApolymerase in its triphosphate form. It has a short half polymerase in its triphosphate form. It has a short half life.life.
REFERENCES:REFERENCES:Daubner, SC et al. Biochemistry. 24:7059Daubner, SC et al. Biochemistry. 24:7059--70657065
Lee L. et al. oligomeric structure of the multifunctional proteinLee L. et al. oligomeric structure of the multifunctional proteinCAD that initiates pyrimidine biosynthesis inCAD that initiates pyrimidine biosynthesis in
mammalian cells. Proc. Nat. Acad of Sci. 1985,mammalian cells. Proc. Nat. Acad of Sci. 1985,82:680282:6802--6806.6806.
Murray, Robert K, et al. Harper¶s review of biochemistry , 25Murray, Robert K, et al. Harper¶s review of biochemistry , 25thth
ed. C. 2000. Appleton and lange. Pp386ed. C. 2000. Appleton and lange. Pp386--401401