Purine and Pyrimidine Metabolism Flashcards
Maybe know the structures of the amino acids
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.
Difference between deoxyribose and ribose
ribose has OH group at 2’ carbon
Starting material for purines
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)
Starting material for pyrimidine
synthesize pyrimidine ring and then attache the ribose-5-phosphate
Rate-limiting step of purine synthesis
Glutamine:phosphoribosyl amidotransferase (or amido phosphoribosyl transferase) Converts PRPP (5-phosphoribosyl-1-pyrophosphate), water, and glutamine into Glutamate, pyrophosphate, and 4-phosphoribosyl 1-amine
What is required for de novo synthesis of a purine?
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
What regulates purine nucleotide synthesis?
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
True or false: De novo synthesis requires less ATP than the salvage pathway.
False
Describe the salvage pathway for purine synthesis
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
Defective protein in Lesch-Nyhan disease?
HGPRT (Hypoxanthine-guanine phosphoribosyl transferase) (Complete lack of protein)
Deficiency of adenosine deaminase can cause?
SCID (Severe combined immunodeficiency)
(No B or T cells)
Causes accumulation of dATP in lymphocytes, which inhibits ribonucleotide reductase
Deficiency of purine nucleoside causes?
Lack of T cells
(No degradation of inosines and guanosines to free bases)
(Nucleosides accumulate)
What would you see an increase of in those with Lesch-Nyhan disease?
Xanthine and Uric acid (the end product of hypoxanthine and guanine)
Regulation of Purine salvage pathway
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.
What disease is related to lack of HGPRTase?
Lesch-Nyhan disease
Description of entirety of Lesch-Nyhan disease
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)
What does allopurinol do?
Inhibits xanthine oxidase, results in buildup up hypoxanthine and xanthine
In de novo pyrimidine synthesis, the carbons and nitrogens come from?
The carbons ( and one nitrogen) come from aspartate and the other nitrogen comes from glutamine amide. One other carbon comes from bicarbonate ion.
Pyrimidine synthesis process
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.
Stuff to know about carbamoyl phosphate synthetase
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)
Hereditary orotic aciduria
results from deficiency in orotate phosphoribosyl transferase or OMP decarboxylase. Treated with uridine (nucleoside, because de novo synthesis not possible).
What makes NDPs?
nucleoside monophosphate kinases (adenylate kinase or guanylate kinase
What protein makes NTPs?
nucleoside diphosphate kinase
True or false: Ribonucleotides are precursors of deoxyribonucleotides.
True
Steps of deoxyribonucleotide synthesis
- Ribonucleotide reductase (rate-limiting)
- Nucleoside 5’-diphosphate kinase
- Deoxycytidylate deaminase
- Thymidylate synthase
- DNA polymerase
True or false: Thymine exists only in deoxy form.
True
True or false: Deoxyribonucleotides are toxic to nonproliferating cells.
True
Things to know about ribonucleotide reductase
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
dTMP synthesis
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
Dihydrofolate Reductase
target for anti-cancer drugs. Converts dihydrofolate to tetrahydrofolate. Aminopterin and methotrexate (amethopterin) inhibit dihydrofolate reductase.
Pyrimidine degradation
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)
Beta-aminoisobutyrate
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
5-fluoracil
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)
Cytosine Arabinoside
Antileukemic, 5’ triphosphate incorporates into DNA, inhibits synthesis of growing DNA strand
Antifolates
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
hydroxyurea
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
Purine analogs as anti-viral agents
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
Pyrimidine analogs as anti-virals
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
Fialuridine
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
