Slideshow is from the University of Michigan Medical School's M1 Renal sequence View additional course materials on Open.Michigan: openmi.ch/med-M1Renal

1. Author(s): Dr. Robert Lyons, 2009
License: Unless otherwise noted, this material is made available under the terms of
the Creative Commons Attribution – Share Alike 3.0 License: http://
creativecommons.org/licenses/by-sa/3.0/
We have reviewed this material in accordance with U.S. Copyright Law and have tried to maximize your ability to use,
share, and adapt it. The citation key on the following slide provides information about how you may share and adapt this
material.
Copyright holders of content included in this material should contact open.michigan@umich.edu with any questions,
corrections, or clarification regarding the use of content.
For more information about how to cite these materials visit http://open.umich.edu/education/about/terms-of-use.
Any medical information in this material is intended to inform and educate and is not a tool for self-diagnosis or a
replacement for medical evaluation, advice, diagnosis or treatment by a healthcare professional. Please speak to your
physician if you have questions about your medical condition.
Viewer discretion is advised: Some medical content is graphic and may not be suitable for all viewers.

2. Citation Key
for more information see: http://open.umich.edu/wiki/CitationPolicy
Use + Share + Adapt
{ Content the copyright holder, author, or law permits you to use, share and adapt. }
Public Domain – Government: Works that are produced by the U.S. Government. (17 USC §105)
Public Domain – Expired: Works that are no longer protected due to an expired copyright term.
Public Domain – Self Dedicated: Works that a copyright holder has dedicated to the public domain.
Creative Commons – Zero Waiver
Creative Commons – Attribution License
Creative Commons – Attribution Share Alike License
Creative Commons – Attribution Noncommercial License
Creative Commons – Attribution Noncommercial Share Alike License
GNU – Free Documentation License
Make Your Own Assessment
{ Content Open.Michigan believes can be used, shared, and adapted because it is ineligible for copyright. }
Public Domain – Ineligible: Works that are ineligible for copyright protection in the U.S. (17 USC § 102(b)) *laws in
your jurisdiction may differ
{ Content Open.Michigan has used under a Fair Use determination. }
Fair Use: Use of works that is determined to be Fair consistent with the U.S. Copyright Act. (17 USC § 107) *laws in your
jurisdiction may differ
Our determination DOES NOT mean that all uses of this 3rd-party content are Fair Uses and we DO NOT guarantee that
your use of the content is Fair.
To use this content you should do your own independent analysis to determine whether or not your use will be Fair.

3. M1 Renal:
Nucleotide Metabolism
Dr. Robert Lyons
Assistant Professor, Biological Chemistry
Director, DNA Sequencing Core
Web: http://seqcore.brcf.med.umich.edu/mcb500
Fall 2008

4. Amino Acid metabolism
Folate metabolism
Amino acids
Met
Methylene Cycle
Glu, Gln, THF
Asp, NH 3
Urea
oxaloacetate
DNA
P u rin e s P y rim id in e s
RNA
Uric Acid (energy)
fumarate
TCA Cycle Nucleic Acid metabolism
R. Lyons

5. Nucleic Acid metabolism
Click on any blue rectangle to see details.
am i n o a c id s ,
Carbamoyl
fo la t e
Phosphate
PRPP
Purine
Salvage Purine Pyrimidine OMP
IM P Biosynthesis Biosynthesis
Pyrimidine
Salvage
P u r in e M P P y r im id in e M P
Ribonucleotide Ribonucleotide
reductase reductase
dNTP DNA dNTP
NTP RNA NTP
Purine Pyrimidine
Degradation Degradation
NH4
U ric A cid ( e n e rg y )
R. Lyons

6. Formation of PRPP: Phosphoribose pyrophosphate
O O
PO P O
OH O P O P
OH OH OH OH
ATP AMP
ribose - 5 - phosphate phosphoribose - 1
R. Lyons
pyrophosphate
PRPP Use in Purine Biosynthesis:
H2
O
O O NH2
PO P O
O PO P
OH OH glutamine glutamate OH OH
phosphoribose - 1
pyrophosphate
R. Lyons

7. O
N
HN
The First Purine: Inosine Monophosphate
N N
(folates are involved in this synthesis)
O
P O
OH OH
R. Lyons
Conversion to Adenosine:
O NH 2
GTP GDP + Pi
N N
HN N
N N N N
aspartate fumarate ribose-5-P
ribose-5-P
Inosine monophosphate Adenosine monophosphate
R. Lyons
Conversion to Guanosine:
H2O
+ + O O
O NAD NADH + H ATP AMP + PPi
N N N
HN N HN
N N O N N NH N N
H 2
gln glu
ribose-5-P ribose-5-P ribose-5-P
Inosine monophosphate Xanthosine monophosphate Guanosine monophosphate
R. Lyons

8. Nucleoside Monophosphate Kinases
AMP + ATP <--> 2ADP
(adenylate kinase)
GMP + ATP <---> GDP + ADP
(guanylate kinase)
• similar enzymes specific for each nucleotide
• no specificity for ribonucleotide vs. deoxyribonucleotide

9. Ribonucleotide Reductase
X
O
P OP O
Enzyme• NADH
OH OH
X
O
P OP O
Enzyme NAD+
OH H
R. Lyons
Hydroxyurea inhibits this enzyme: chemotherapeutic use
O
HONH
C
NH
2

10. Regulation of Ribonucleotide Reductase
ATP
CDP + dCDP dCTP
(-)
ATP
UDP + dUDP dTTP
(-)
GDP + dGDP dGTP
ADP + dADP dATP
(-)
R. Lyons

11. Nucleoside Diphosphate Kinase
N1DP + N2TP <--> N1TP + N2DP
dN1DP + N2TP <--> dN1TP + N2DP
• No specificity for base
• No specificity for ribo vs deoxy

12. Feed-forward regulation by PRPP
PRPP
IMP
ATP GTP
GMP AMP
GTP ATP
R. Lyons

13. Feed-forward regulation by PRPP
PRPP
+
IMP
ATP GTP
GMP AMP
GTP ATP
R. Lyons

14. Feed-forward regulation by PRPP
PRPP
IMP
ATP GTP
+ +
GMP AMP
GTP ATP
R. Lyons

15. Feed-forward regulation by PRPP
PRPP
+
- -
IMP
- -
ATP GTP
GMP AMP
GTP ATP
R. Lyons

16. Degradation of the Purine Nucleosides:
+
NH 2 HO NH O Pi ribose - 1 - P O
2 4
N N N
N HN HN
Adenosine purine nucleoside
N N N N N N
deaminase phosphorylase H
ribose (ADA) ribose
Adenosine Inosine Hypoxanthine
xanthine
oxidase
Pi ribose - 1 - P
O + O
O H 2O NH 4
N N
HN N HN
HN
N purine nucleoside NH Guanine O N N
NH 2 N N N H H
phosphorylase 2
H deaminase
ribose
Xanthine
Guanosine Guanine xanthine
oxidase
O
H
N
HN
Uric acid O
O N N
H H
R. Lyons

17. Salvage Pathways for Purine Nucleotides
O
N O
HN
Hypoxanthine- N
N N guanine HN
H
Hypoxanthine phosphoribosyl
N N
transferase
+ + PPi
O
P O
O
P O
O P O P OH OH
OH OH Inosine monophosphate
PRPP
R. Lyons
APRT - Adenine phosphoribosyl transferase -
performs a similar function with adenine.

18. Adenosine Deaminase Deficiency:
NH 2
N O O
N
N N
HN HN
N N
N N N N
HO O Adenosine H
deaminase 2-deoxyribose
(ADA) Hypoxanthine
Deoxyinosine
OH H
Deoxyadenosine
Guanine Xanthine
dAMP
dADP
Uric acid
dATP
R. Lyons

19. O
HN
N Gout: deposition of urate crystals in joints,
N N tophi in cooler periphery
H
Hypoxanthine
xanthine Hyperuricemia can be caused by:
oxidase
O Accelerated degradation of purines:
HN
N
O N
H
N
H
•Accelerated synthesis of purines
Xanthine
•Increased dietary intake of purines
xanthine
oxidase
Impaired renal clearance of uric acid
O
H
N OH
HN H
O N
N
O N
H
N
H
Allopurinol inhibits xanthine oxidase
N
N
Uric acid and reduces blood uric acid levels:
Allopurinol
R. Lyons
R. Lyons

20. An 80-year-old man with a 30-year
history of gout, this patient had been
treated intermittently to reduce his
serum urate levels.
The New England Journal of Medicine

21. Lesch-Nyhan Syndrome: Defective HGPRT
• hyperuricemia
• spasticity
• mental retardation
• self-mutilation behavior
O
N O
HN
Hypoxanthine- N
N N guanine HN
H
Hypoxanthine phosphoribosyl
N N
transferase
+ + PPi
O
P O
O
P O
O P O P OH OH
OH OH Inosine monophosphate
PRPP
R. Lyons
A defect in APRT does NOT have similar consequences

22. Myoadenylate Deaminase Fills the TCA Cycle in Muscle
H 2O NH 3
myoadenylate
deaminase
NH 2
O
N
N N
HN
N N
N N
ribose-5-P ribose-5-P
Adenosine monophosphate Inosine monophosphate
Asp, GTP
Fumarate
GDP + Pi
To
TCA
Cycle
R. Lyons

24. Carbamoyl phosphate synthetase II - a cytoplasmic enzyme…
O
- 2-
2ATP + HCO3 + + glutamate + 2ADP + Pi
NH2 C OP
glutamine + H 2O
carbamoyl
phosphate
R. Lyons
…used for pyrimidine synthesis
O O
- O
O
NH2 -
O C C
CH2 NH2 CH2 HN
C 2-
+
O OP CH C CH O
- - N CO2
NH3 CO2 O N CO2 H
+
carbamoyl H
phosphate aspartate orotate
R. Lyons

25. Orotate is linked to PRPP to form Uridine monophosphate:
O
HN
O O
O N CO2
H HN HN
orotate
O N CO2 O N
+ O
P O O
P O
O
P O CO2
O P OP OH OH OH OH
OH OH orotidine uridine
phosphoribose - 1 monophosphate monophosphate
pyrophosphate
R. Lyons

26. Newly-synthesized uridine monophosphate will be phosphorylated to
UDP and UTP, as described for the purine nucleotides.
UTP can be converted to CTP by CTP Synthetase:
O NH 2
HN N
O N gln glu O N
O O
P OP O P O P OP O P O
OH OH ATP ADP OH OH
+ +
uridine H 2O Pi cytidine
triphosphate triphosphate
R. Lyons

27. Some UDP is converted to dUDP via ribonucleotide reductase.
UDP dUDP dUTP dUMP
ribo-
nucleotide
reductase ATP ADP H 2O Pi
R. Lyons
The Thymidylate Synthase Reaction:
O O
HN HN CH3
O N O N
O O
P O P O
thymidylate synthase
OH H OH H
deoxyuridine 5 10 deoxythymidine
monophosphate N ,N
methylene monophosphate
dihydrofolate
tetrahydrofolate
DHFR NADH
****
methylene THF
NAD+
donor
R. Lyons

28. Methotrexate Inhibits Dihydrofolate Reductase:
O O
HN HN CH3
O N O N
O O
P O P O
thymidylate synthase
OH H OH H
deoxyuridine deoxythymidine
monophosphate N 5, N10
methylene monophosphate
dihydrofolate
tetrahydrofolate
DHFR NADH
**** X Methotrexate
methylene THF
donor NAD+
R. Lyons
Dihydrofolate builds up, levels of THF become limiting,
thymidylate synthase is unable to proceed. Follow it with
a dose of Leucovorin, a.k.a. formyl-THF.

29. FdUMP Inhibits The Thymidylate Synthase Reaction:
O O
HN F HN CH3
O N O N
O O
P O P O
thymidylate synthase
OH H
X
OH H
5-fluorodeoxyuridine deoxythymidine
monophosphate N 5, N10
monophosphate
(F-dUMP) methylene dihydrofolate
tetrahydrofolate
DHFR NADH
****
methylene THF
NAD+
donor
R. Lyons

30. Complicated Pathways for Pyrimidine Production:
dCTP CTP UTP dUTP dTTP
***
dCDP CDP UDP dUDP dTDP
*** ***
UMP dUMP dTMP
***
de-novo synthesis OMP
R. Lyons
This figure is primarily a study aid; you do not need to memorize it or reproduce it.
The information here merely summarizes material from previous sections.

31. Pathologies of pyrimidine nucleotide biosynthesis:
Orotic acidurea due to OTC deficiency - please
review your Urea Cycle notes.
Hereditary orotic acidurea - deficiency of the
enzyme that convert orotate to OMP to UMP.
Not common.

32. Pyrimidine degradation:
Cytidine deaminase converts cytidine to uridine
NH2 O
HN
O
N
N O N
cytidine
HO
O deaminase HO
O
OH OH OH OH
Cytidine Uridine
R. Lyons
A phosphorylase removes the sugar
O
O HN CH3
O
HN
N
CH3
Pi
pyrimidine
phosphorylase
O N
H
thymine
+
HO
O
HO
+
O OP
OH H
(deoxy)thymidine OH H
deoxyribose-1-phosphate
R. Lyons
Degradation of the base proceeds (products are
unimportant here)

33. Pyrimidines can be salvaged as well:
Enzyme: Pyrimidine nucleoside phosphorylases
Thymine + deoxyribose-1-phosphate --> thymidine
(NOT thymidine monophosphate!)
O
O HN CH3
HN CH3 O N
pyrimidine H
O N Pi phosphorylase thymine
+
O +
HO
O OP
HO
OH H
(deoxy)thymidine OH H
deoxyribose-1-phosphate
R. Lyons
Enzyme: Thymidine kinase - adds the monophosphate back
Thymidine + ATP --> thymidine monophosphate
Herpes Simplex Virus carries its own tk gene

34. Certain drugs act via the pyrimidine salvage pathway:
G HSV G
HO O thymidine P O O
kinase
H H H H
Acyclovir AcycloGMP
R. Lyons
O
HN F O
O
HN F
O N HN F
H pyrimidine O N uridine
5-fluorouracil phosphorylase kinase O N
O
+ HO O
O P O
HO OP
OH H
OH H
OH H
deoxyribose-1-phosphate fluorodeoxyuridine fluorodeoxyuridine monophosphate
(FdUMP)
R. Lyons

35. 5-FU efficacy depends on rate of degradation vs activation
O
HN F O
O
HN F
O N HN F
H pyrimidine O N uridine
5-fluorouracil phosphorylase kinase O N
O
+ HO O
O P O
HO OP
OH H
OH H
OH H
deoxyribose-1-phosphate fluorodeoxyuridine fluorodeoxyuridine monophosphate
(FdUMP)
R. Lyons
5-FU --> --> FdUMP
+ methylene-THF + Thymidylate Synthase
--> inactivation of TS
Degradation
(via dihydropyrimidine dehydrogenase, DPD
DPD inhibitors can potentiate 5FU activity

36. Capecitabine mode of action:
O
CH3
O
HN
N F
O
O N cytidine pyrimidine
dealkylation deaminase phoshorylase
HN F
O
HO O N
H
OH OH fluorouracil
capecitabine
R. Lyons
Cytosine arabinoside (araC) activation and inactivation:
O NH2 NH2
HN
N N
O N
cytidine O N cytidine O N
O deaminase O kinase
HO O
OH HO P O
OH OH
OH OH OH
Uridine arabinoside Cytosine arabinoside (araC) Cytosine arabinoside monophosphate
(INACTIVE) (ACTIVE)
R. Lyons

37. Additional Source Information
for more information see: http://open.umich.edu/wiki/CitationPolicy
Slide 4: Robert Lyons
Slide 5: Robert Lyons
Slide 6: Robert Lyons
Slide 7: Robert Lyons
Slide 9: Robert Lyons
Slide 10: Robert Lyons
Slide 12: Robert Lyons
Slide 13: Robert Lyons
Slide 14: Robert Lyons
Slide 15: Robert Lyons
Slide 16: Robert Lyons
Slide 17: Robert Lyons
Slide 18: Robert Lyons
Slide 19: Robert Lyons
Slide 20: The New England Journal of Medicine, http://content.nejm.org/cgi/content/full/353/23/e20
Slide 21: Robert Lyons
Slide 22: Robert Lyons
Slide 24: Robert Lyons
Slide 25: Robert Lyons
Slide 26: Robert Lyons
Slide 27: Robert Lyons
Slide 28: Robert Lyons
Slide 30: Robert Lyons
Slide 32: Robert Lyons
Slide 33: Robert Lyons
Slide 34: Robert Lyons
Slide 35: Robert Lyons
Slide 36: Robert Lyons