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【目的与要求】• 记住嘌呤核苷酸有两条合成途径。结合嘌呤核苷酸
结构与从头合成途径,说出嘌呤核苷酸各元素或组件的材料来源。熟记二磷酸核苷还原生成脱氧嘌呤核苷酸。写出与嘌呤核苷酸补救合成有关的酶的名称、功能、酶缺陷相关的疾病
• 结合嘌呤核苷酸合成途径、调节,熟记嘌呤核苷酸抗代谢药物作用机理及临床意义
• 记住嘌呤核苷酸体内分解代谢终产物 -尿酸及其与医学的关系
• 熟记嘧啶核苷酸从头合成的原料及合成调节。说出嘧啶核苷酸补救合成所需的酶及其催化的反应。明白嘧啶核苷酸抗代谢药物作用机理,记住嘧啶核苷酸分解代谢产物名称
• 8.1 Purine metabolism
-8.1.1 The Biosynthesis of Purines
-8.1.2 Purine Salvage
-8.1.3 De-oxyribonucleotide Synthesis
-8.1.4 Purine Degradation
• 8.2 Pyrimidine metabolism
-8.2.1 Biosynthesis of Pyrimidines
-8.2.2 Pyrimidine Degradation
Outline
Deoxyribonucleotide umol
food
protein nuclear acid (RNA and DNA) (intestine)
Endonucleases(phosphodiesterase)
mononucleotide
Nucleotidase(phosphoesterase)
nucleosidePhosphate
nucleosidase
Ribose or ribose-1-phosphatebase
Nuclear acid digestion
Uric acid (purines)
β-ureidopropionate( primidines)
RNase
DNase
(stomach)
ribonucleotide mmol
( 戊糖代谢 )excrete
Biological Roles of Nucleotides
• Monomeric units of nucleic acids *• “ Energy currency”(ATP) *• Regulation of physiological processes
– Adenosine controls coronary( 冠脉 ) blood flow– cAMP and cGMP serve as signaling molecules
• Precursor function - GTP to tetrahydrobiopternin• Coenzyme components - 5’-AMP in FAD/NAD+• Activated intermediates: UDP-Glucose• Allosteric effectors- regulate themselves and others
8.1.1 Nucleotide Biosynthesis• For both purines and pyrimidines there are
two means of synthesis
- de novo (from bits and parts)
- salvage (recycle from pre-existing nucleosides,and bases)
• Ribose generates energy, but purine and pyrimidine rings do not
• Nucleotide synthesis pathways are good targets for anti-cancer/antibacterial strategies
Bases/Nucleosides/Nucleotides
BaseBase + Sugar=
NucleosideBase + Sugar + Phosphate=
NucleotideAdenine Deoxyadenosine Deoxyadenosine
5’-triphosphate(dATP)
De novo purine biosynthesis
• John Buchanan (1948) "traced" the sources of all ni
ne atoms of purine ring
1. In de novo synthesis, Inosine-5'-P (Inosine Monophosphate, IMP) is the first nucleotide formed
2. It is ,then, converted to either AMP or GMP Location: liver cellular CytoplasmCytoplasm
De novo purinenucleotide synthesis proceeds by the synth
esis of the purine base upon the ribose sugar moiety
The metabolic origin of the nine atoms The metabolic origin of the nine atoms in the purine ring systemin the purine ring system
•N-1: aspartic acid •C-2:THF - one carbon units•N-3: glutamine •C-4, C-5, N-7: glycine •C-6: CO2 •C-8: THF - one carbon units•N-9: glutamine
N-1
C-2
N-3
C-6
C-8
N-9
H
甘氨当中站 , 谷氮坐两边 ,
左上天冬氨 , 头顶 CO2
还有俩一碳
N-7
C-4
C-5
R-5'-P
PRPP synthetase
PP-1'-R-5'-P(PRPP)
OH
O
OH P O
OH
H
OH
H
OH
HH
O
OH
O
OH P O
OH
H
OH
H
OH
HH
O
ATP
P_ P
NH3 via glutamine
NH3 via glutamine
1 carbon via folate
1 carbon via folate
NH3 via aspartyl- succinate
(2) ATP dependent step
ATP dependent step
ATP dependent step
ATP dependent step
ATP dependent step
PRPP
Inosine monophosphate
Regulation of De Novo Synthesis
ATP provides the energy for GMP synthesis
GTP provides the energy for AMP synthesis
3. “cross regulation” occurs from IMP to AMP and GMP
2.End product inhibition and “feed forward” regulation
1. Committed Steps( at the first two steps ):
PRPP , PRA(A bunch of steps you don’t need to know)
Purines are synthesized on the Ribose ring
FeedbackInhibition
8.1.2 Salvage Pathway for Purines
Hypoxanthineor
Guanine
+ PRPP = IMP or GMP + PPi Hypoxanthineguanosylphosphoribosyl transferase
(HGPRTase)
Adenine + PRPP = AMP + PPi Adeninephosphoribosyl transferase
(APRTase)
Salvage pathways are particularly important in brain/marrow that lack de novo purine synthesis
Lesch-Nyhan Syndrome( 莱 - 尼综合症 )
Absence of HGPRTaseX-linked (Gene on X)
Occurs primarily in malesCharacterized by:
purine synthesis is increased 200-foldIncreased uric acidSpasticity( 痉挛 )Neurological defectsAggressive behaviorSelf-mutilation( 自残 )
AMP GMP
XMP AMPS(腺苷酸代琥珀酸)
IMP
NH3
Adenine DeaminaseNADP
+NH 3
NADPH
Guanine Red
uctase
Inter-conversion of Purine nucleotides
OH
HHO
H
H
HOCH2 OH
H
1´
2´3´
4´
5´BASE
OH
HHO
H
H
HOCH2
OH
OH
1´
2´3´
4´5´
BASE
Deoxyribonucleoside Ribonucleoside
RibonucleotideReductase
8.1.3 Deoxyribonucleotide Biosynthesis
NDP dNDPribonucleotide reductase
ADP dADP
ribonucleotide reductase
GDP dGDP
ribonucleotide reductase
UDP dUDP
ribonucleotide reductase
CDP dCDP
ribonucleotide reductase
TDP dTDP
Deoxyribonucleotide Biosynthesis ?
Mg2+硫氧还蛋白
Ribonucleotides can be converted to deoxyribonucleotides
by Ribonucleotide Reductase at the diphosphate level
The ribonucleotide reductase, An (R1)2(R2)2- type enzyme
, has R1 (86 kD) and R2 (43.5 kD) two subunits
E. coli Ribonucleotide Reductase
Regulates the level of
cellular dNTPs
dNDP+ATP dNTP+ADPkinase
kinasedCDP+ATP dCTP+ADP
dUDP+ATP dUTP+ADPkinase
dGDP+ATP dGTP+ADPkinase
dADP+ATP dATP +ADPkinase
dTTP ?
dNDP dNMP+Piphosphorylase
Regulation of dNTP Synthesis
• The overall activity of ribonucleotide reductase must be regulated
• Balance of the four deoxynucleotides must be controlled
• ATP activates, dATP inhibits at the overall activity site
• ATP, dATP, dTTP and dGTP bind at the specificity site to regulate the selection of substrates and the products made
over-growth + Heterogeneity
( nucleotides + protein )
Tumor
How to inhibit the biosynthesis of the tumor cells?
for anti-cancer strategies(antibacterial)
Chemotherapeutic Agents1. Analogs of purine:
8- 氮杂鸟嘌呤
N
OH
N
NNH
N
SH
N
NNH
N
SH
N
NNH
H2N
N
OH
N
NNH
N
6- 巯基鸟嘌呤
inosine6- 巯基嘌呤
( 6-mercaptopurine,
6-MP )
(8-azoguanine)
(6-mercaptoguanine)
2. Analogs of amino acids:
H2N—C—CH2—CH2—CH—COOH
O NH2
N+ —N—CH2—C—O—CH2—CH—COOH
O NH2
N+ —N—CH2—C—CH2—CH2—CH—COOH
O NH2
Gln
氮杂丝氨酸( azaserine )
6- 重氮 -5- 氧正亮氨酸 (diazonnorleucine)
Inhibit the reactions
of the Gln
3. Analogs of Folic acid
R=H , aminopterin, 氨喋呤R=CH3 , methotrexate,
氨甲喋呤 ,MTX
N
NH2
N
NNH
H2N
—CH2—N—
R
—C—N—CH
O
CH2
CH2
COOH
COOHH
N
NCH
—CH2—N—
H
—C—N—CH
O
CH2
CH2
COOH
COOHH
H2N
N
OH
NH
四氢叶酸 ,THF
PRPP
Gln
6MP
氮杂丝氨酸 (azaserine)
PRA GAR FGAR
FGAM
MTX
azaserine
AICAR
MTXFAICAR IMP
AMP
GMP
PRPPPPi
PPi PRPP
6MP
6MP
6MPazaserine
A
IG
PRPPPPiThe mechanism of the Chemotherapeutic Agents
AMP I
GMP G
XXO
XO
Excreted in
Urine
Sequential removal of bits and pieces
End product is uric acid
XO: Xanthine Oxidase
8.1.4 Purine catabolismcatabolism
The scale of uric acid (normal value) :
0.12 ~ 0.36mmol/L;
male, 0.27mmol/L;
formale, 0.21mmol/L
> 0.48mmol/L(8mg%),
Xanthine Oxidase and Gout
C
OH
N
NNH
H
NN
OH
N
NNH
CH
I
allopurinol
PRPP
Allopurinol nucleotideXO
Purine nucleotides
The mechanism of allopurinol as a treatment of gout
↓
Uric acids
X
8.2 Pyrimidine Biosynthesis
Pyrimidine Biosynthesis:
In contrast to purines, First, synthesis of
the pyrimidine ring; Next, attachment of ribos
e-phosphate to the ring
Carbamoyl-PAspartate
De Novo Pyrimidine Biosynthesis
The metabolic origin of the six atoms The metabolic origin of the six atoms of the pyrimidine ringof the pyrimidine ring
NH2HCH2C
HOOC
HOOC
1. Thymine nucleotides are made from dU
MP, which derives from dUDP, dCDP
2. Biosynthesis of deoxyribonucleotides b
y ribonucleotide reductase
3. Biosynthesis of thymidine monophosphate (dTMP)
by thymidylate synthase
Synthesis of Thymine Nucleotides
CO
HNC
CH
CHN
O
dR-5'-P
dUMP
CO
HNC
C-CH3
CHN
O
dR-5'-P
dTMP synthase
FH2N5,N10- methylene FH4
FH4
DHFR
NADPH+H+NADP+
dTMP
kinasedTDP
kinase
ADP
dTTP
ATP ADP
dTMPATP
Thymidylate synthase methylates dUMP at 5-position to make dTMP
N5,N10-methylene THF is 1-C donor
Regulation of Pyrimidine Synthesis ( de novo )
• Aspartate transcarbamoylase (ATCase ,细菌 ) catalyzes the condensation of carbamoyl
phosphate with aspartate to form carbamoyl-a
spartate
• Note that carbamoyl phosphate represents an
‘activated’ carbamoyl group
Feedback Inhibition
Regulation of Pyrimidines Biosynthesis
Regulation occurs at first step in the pathway (committed step)
Inhibited by UTPIf you have lots of UTP around this means you won’t
make more that you don’t need
×2ATP + CO2 + Glutamine = carbamoyl phosphate
CPS II
• Carbamoyl phosphate for pyrimidine synthesis is made by carbamoyl phosphate synthetase II (CPS II ,哺乳动物细胞 )
• This is a cytosolic enzyme (whereas CPS I is mitochondrial and used for the urea cycle)
• Substrates are HCO3-, glutamine, 2 ATP
Allosteric regulation of pyrimidine biosynthesis
Enzyme regulated Allosteric effector Effect
carbamoyl phosphate synthetase II
UDP, UTP Feedback inhibition
PRPP, ATP stimulatory
Biosynthesis: Purine vs. Pyrimidine
start with ribose, build on nitrogen baseRegulated by GTP/ATPGenerates IMPRequires Energy
build nitrogen base then added to PRPPSynthesized Regulated by UTPGenerates UMP/CMPRequires Energy
“Both are very complicated multi-step process whichyour kindly professor does not expect you to know in detail”
Salvaging Pyrimidines
• Pyrimidines+PRPP Nucleoside+PPi ( 嘧啶磷酸核糖转移酶 )• A second type of salvage pathway involves two steps a
nd is the major pathway for the pyrimidines, uracil and thymine
Base + Ribose 1-phosphate = Nucleoside + Pi (nucleoside phosphorylase)
• Nucleoside + ATP Nucleotide + ADP (nucleoside kinase-irreversible)
Analogs of pymidines /pymidine nucleosides:
5- 氟尿嘧啶 ,5-Fu
阿糖胞苷 Cytarabine
环胞苷Cyclocytidine
N
NH
O
O
F
HHOH2C
H
HHO
OH
HH
O
C
C
C
N
N
CO
NH2
HOH2C
H
H
OH
HH
O
C
C
C
N
N
C
NH·HCl
O
Inhibitors of pymidines synthesis are cancer drugs
CO
NC
CH
CHN
NH2
HC
O
HNC
CH
CHN
O
H
NH3
CO
HNC
CH2
CH2N
O
H
NADPH+H+ NADP+
H2O
CO
H2NC
CH2
CH2N
O
H
HOH2OCO2+NH3
H2N-CH2-CH2-COOH
β-Alanine
C U
Pyrimidine Catabolism-1
CO
HNC
C-CH3
CHN
O
H
NADPH+H+ NADP+
CO
HNC
CH-CH3
CH2N
O
H
CO
H2NC
CH-CH3
CH2N
O
H
HOH2OCO2+NH3
β-aminoisobutyrate
H2N-CH2-CH-COOH
CH3
β- 脲基异丁酸
H2OT DHT
Pyrimidine Catabolism-2
β- 氨基异丁酸
overview5'-P-R
PRPP
IMP
dAMP GMPdGMPAMP
dADP GDPdGDPADP
dATP GTPdGTPATP
UMP CMPdUMP
UDP CDPdUDP
UTP CTP
dUTP
dTMPdCMP
dTDPdCDP
dTTPdCTP
CO2+Gln
H2N-CO-P
OMP
De novo synthesis
dUDP
dCMPdUMP
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