Purine and Pyrimidine Metabolism

Question 1

Maybe know the structures of the amino acids

Answer 1

Actually yeah know them
Guanine and Adenine are purines. Adenine has an amine on carbon 6. Guanine has a ketone on carbon 6 (and an amine on carbon 2). The other ring is five carbons (three more).
Cytosine, Uracil, and Thymine are pyrimidines. Uracil has two ketones in the carbon 2 and 4 places. Cytosine has a ketone at carbon 2 and an amine at carbon 4. Thymine is basically uracil (ketone at carbons 2 and 4) with a methyl group at carbon 5.

Question 2

Difference between deoxyribose and ribose

Answer 2

ribose has OH group at 2’ carbon

Question 3

Starting material for purines

Answer 3

ribose-5-phosphate (build on the purine base)

Glycine, 2 Glutamine amides, and an aspartate amino combined with bicarbonate, and two C1-H4folate. Requires 6 ATP and 5-phosphoribosyl-1-pyrophosphate (PRPP)

Question 4

Starting material for pyrimidine

Answer 4

synthesize pyrimidine ring and then attache the ribose-5-phosphate

Question 5

Rate-limiting step of purine synthesis

Answer 5

Glutamine:phosphoribosyl amidotransferase (or amido phosphoribosyl transferase)
Converts PRPP (5-phosphoribosyl-1-pyrophosphate), water, and glutamine into Glutamate, pyrophosphate, and 4-phosphoribosyl 1-amine

Question 6

What is required for de novo synthesis of a purine?

Answer 6

Glutamine and PRPP become 5’-phosphoribosylamine
ATP and Glycine
Formyl-THF
ATP and Glutamine
ATP
CO2
ATP and Aspartate
Formyl-THF
(So 4 ATP, 2 Formyl-THF, Glycine, Glutamine, Aspartate, and CO2 forms IMP)
Then GTP and Aspartate is used to make AMP
and ATP and Glutamine is used to make GMP

Question 7

What regulates purine nucleotide synthesis?

Answer 7

PRPP synthetase is inhibited by ADP and GDP
Glutamine phosphoribosyl amido transferase is inhibited by GMP and AMP
IMP dehydrogenase is inhibited by GMP
Adenylosuccinate synthetase is inhibited by AMP

Question 8

True or false: De novo synthesis requires less ATP than the salvage pathway.

Answer 8

False

Question 9

Describe the salvage pathway for purine synthesis

Answer 9

There are two pathways
There is a hypoxanthine-guanine phosphoribosyl transferase pathway (HGPRTase)
(It uses hypoxanthine or guanine and PRPP (5-phosphoribosyl-1-pyrophosphate)
(Makes GMP or IMP and pyrophosphate)

The other pathway uses adenine phosphoribosyltransferase (APRTase), which uses adenine and PRPP to make AMP and pyrophosphate

Question 10

Defective protein in Lesch-Nyhan disease?

Answer 10

HGPRT (Hypoxanthine-guanine phosphoribosyl transferase) (Complete lack of protein)

Question 11

Deficiency of adenosine deaminase can cause?

Answer 11

SCID (Severe combined immunodeficiency)
(No B or T cells)
Causes accumulation of dATP in lymphocytes, which inhibits ribonucleotide reductase

Question 12

Deficiency of purine nucleoside causes?

Answer 12

Lack of T cells
(No degradation of inosines and guanosines to free bases)
(Nucleosides accumulate)

Question 13

What would you see an increase of in those with Lesch-Nyhan disease?

Answer 13

Xanthine and Uric acid (the end product of hypoxanthine and guanine)

Question 14

Regulation of Purine salvage pathway

Answer 14

HGRPTase inhibited by IMP and GMP. APRTase inhibited by AMP

AMP and GMP also inhibits de novo synthesis pathway. The use of PRPP also reduces de novo synthesis pathway.

Question 15

What disease is related to lack of HGPRTase?

Answer 15

Lesch-Nyhan disease

Question 16

Description of entirety of Lesch-Nyhan disease

Answer 16

Complete lack of HGPRT
X-linked (males affected)
No salvage pathway, increased uric acid, greater de novo purine synthesis
Severe neurological disease and urate nephrolithiasis (kidney damage)
Hyperuricemia

Left untreated, death in first decade. Treated with allopurinol (competitive inhibitor of xanthine oxidase) (allopurinol prevents nephropathy but does not affect neurological or behavior symptoms)

Question 17

What does allopurinol do?

Answer 17

Inhibits xanthine oxidase, results in buildup up hypoxanthine and xanthine

Question 18

In de novo pyrimidine synthesis, the carbons and nitrogens come from?

Answer 18

The carbons ( and one nitrogen) come from aspartate and the other nitrogen comes from glutamine amide. One other carbon comes from bicarbonate ion.

Question 19

Pyrimidine synthesis process

Answer 19

nitrogenous base assembled, then sugar and phosphate added. Nitrogenous base comes from aspartate, glutamine, and bicarbonate ion

carbamoyl phosphate synthetase takes glutamine, carbon dioxide, and two ATP to create 2 ADP, glutamate, phosphate, and carbamoyl phosphate. That is converted to N-carbamoyl aspartate, then hihydroorotate, orotate, and then OMP (Orotidine 5’-mono-phosphate). Orotidine 5’-mono-phosphate decarboxylase converts it to UMP, which is converted to UTP and then CTP.

Question 20

Stuff to know about carbamoyl phosphate synthetase

Answer 20

CPSII is cytoplasmic. It uses nitrogen from glutamine. It is activated by PRPP (5-phosphoribosyl-1-pyrophosphate). It is inhibited by UTP

(CPSI is in mitochondria and uses ammonia as a an amino acid source)

Question 21

Hereditary orotic aciduria

Answer 21

results from deficiency in orotate phosphoribosyl transferase or OMP decarboxylase. Treated with uridine (nucleoside, because de novo synthesis not possible).

Question 22

What makes NDPs?

Answer 22

nucleoside monophosphate kinases (adenylate kinase or guanylate kinase

Question 23

What protein makes NTPs?

Answer 23

nucleoside diphosphate kinase

Question 24

True or false: Ribonucleotides are precursors of deoxyribonucleotides.

Answer 24

True

Question 25

Steps of deoxyribonucleotide synthesis

Answer 25

1. Ribonucleotide reductase (rate-limiting)
2. Nucleoside 5’-diphosphate kinase
3. Deoxycytidylate deaminase
4. Thymidylate synthase
5. DNA polymerase

Question 26

True or false: Thymine exists only in deoxy form.

Answer 26

True

Question 27

True or false: Deoxyribonucleotides are toxic to nonproliferating cells.

Answer 27

True

Question 28

Things to know about ribonucleotide reductase

Answer 28

Requires thioredoxin, reduces all four NDPs to dNDPs

Two allosteric sites, one controls activity, one controls specificity

ATP activates enzyme, dATP inhibits enzyme

Substrate specificity regulation is complex. Effector site binding affects preferred substrate in an effort to balance production of all four dNDPs

Question 29

dTMP synthesis

Answer 29

from dUMP, FH4 is the single carbon donor and a reducing agent

Thymidylate synthase converts dUMP to dTMP using FH4 (N5,N10-tetrahydrofolate)

Inhibited by fluoracil

Question 30

Dihydrofolate Reductase

Answer 30

target for anti-cancer drugs. Converts dihydrofolate to tetrahydrofolate. Aminopterin and methotrexate (amethopterin) inhibit dihydrofolate reductase.

Question 31

Pyrimidine degradation

Answer 31

Uracil is degraded to beta-alanine. (converted to dihydrouracil and beta-ureidopropionate first)
Thymine is degraded to beta-aminoisobutyrate (converted to dihydrothymine and beta-ureidoisobutyrate first)

Question 32

Beta-aminoisobutyrate

Answer 32

excreted in urine, exclusively from thymine, measure to estimate DNA turnover, increases in cancer patients receiving chemotherapy or radiation due to cell death and DNA degradation

Question 33

5-fluoracil

Answer 33

Chemotherapeutic agent. Treats solid tumors.
Active species: 5-fluoridine 5’triphosphate FUTP (incorporated into RNA, inhibits maturation of 48S precursor rRNA into 28S and 18S rRNA)
FdUMP (potent inhibitor of thymidylate synthase (inhibits dUMP to dTMP synthesis), dUMP is incorporated into DNA and attempts to remove and repair causes DNA breaks)

Question 34

Cytosine Arabinoside

Answer 34

Antileukemic, 5’ triphosphate incorporates into DNA, inhibits synthesis of growing DNA strand

Question 35

Antifolates

Answer 35

Inhibit tetrahydrofolate formation

methotrexate treats human cancers, inhibits dihydrofolate reductase. Toxic unless supplemented with thymidine and hypoxanthine

In leukemia treatment, normal cells are rescued from the toxic effects of high-dose methotrexate by N5-formyl-FH4 (leucovorin), increasing clinical efficacy of methotrexate treatment

Question 36

hydroxyurea

Answer 36

specific inhibition of DNA synthesis without inhibiting RNA or protein synthesis
Acts on ribonucleotide reductase
Limited clinical use due to rapid rate of clearance and the high drug concentrations required for effective inhibition

Question 37

Purine analogs as anti-viral agents

Answer 37

acyclovir:
acycloguanosine (purine analog)
activated to the monophosphate by a viral kinase (HSV-thymidine kinase) (mamalian thymidine kinase can’t use as a substrate)
The triphosphate form is a substrate for the HSV-specific DNA polymerase (Causes DNA chain termination of viral gene products)

Ganciclovir (purine analog)
same mechanism as acyclovir, used to treat cytomegalovirus infections in HIV patients

Question 38

Pyrimidine analogs as anti-virals

Answer 38

AZT: Azido-d’-deoxythymidine (pyrimidine analog)
phosphorylated by cellular kinases
Blocks HIV replication by inhibiting HIV-DNA polymerase (an RNA-dependent polymerase)
The HIV polymerase is >100x more sensitive than host

DDI (2’,3’-dideoxyinosine) - another pyrimidine analog with chain termination mechanism

Question 39

Fialuridine

Answer 39

FIAU 1-(2-deoxy-2-fluoro-arabinosyl)-5-iodouracil was developed as an anti-viral against hepatitis B virus
5/15 patients died of liver failure in clinical trial
Incorporated into mitochondrial DNA via DNA polymerase gamma, prevents mitochondrial division
Lactic acidosis due to increased anaerobic glycolysis
No side effects detected in animal studies
Led to analysis of clinical trials rules