BB 451 Nucleotide Metabolism
NUCLEOTIDE METABOLISM
Why is it important to balance amounts of nucleotides?
imbalances can lead to mutation
Contrast nucleoside with nucleotide vs nucleic acid
nucleoside:sugar and a base
nucleotide: base, sugar, and at least one phosphate
nucleic acid: made up of many nucleotides
What is a “base”?
a nitrogen-containing organic molecule, specifically a purine or pyrimidine
What are the sugars in DNA and RNA?
RNA: ribose
DNA: Deoxyribose
What is meant by de novo and salvage?
de novo: ribonucleotides are made from scratch
Salvage: ribonucleotides are made from recycled parts (“cheaper’)
Which are the purines? Pyrimidines?
purines: Adenine (A) and Guanine (G) are the purines.2 carbon rings
made from glycine, glutamine, aspartate, carbon dioxide, and folate derivativesmade with ring on ribose. inosine is ring structure
Pyrimidines: Cytosine (C), Thymine (T), and Uracil (U) are the pyrimidines.
1 carbon rings.made from aspartate and glutamine with the addition of carbon dioxide
ring and ribose are made separate, orotate is ring structure
What are the starting components for the bases?
amino acids, one carbon donors, co2
Is it surprising that DNA and RNA formed on early Earth? Why or why not?
No, building blocks were readily available
What is important about ribose-5-phosphate?
starting molecule for synthesis of purines
What is PRPP? Is it for purines or pyrimidines or both?
used in both. Ribose with 1 phosphate of one end and 2 on the other
What is the significance of inosine monophosphate?
IMP, branch molecules. split into either adenine-containing or guanine-containing
How is de novo synthesis of AMP and GMP balanced?
Through negative feedback. They inhibit PRPP amidotransferase. balance b/w two maintained because both need to bind to completely shut off. If only one shut off wouldn't be able to make the other. can regulate own pathways so one is made instead of the other
How is PRPP amidotransferase regulated?
turned off by both AMP and GMP binding to it (feedback inhibitors)
How is PRPP synthase inhibited?
by high phosphate and ADP
Are phosphates put on the bases one at a time or all three at once?
One at a time
What is the source of energy to make guanine nucleotides? How about vice versa? Why does this work?
ATP for guanine. GTP for adenine. good that use other as energy because it wouldn't work to have to use something as a source to make something because you already don't have it
When converting a nucleoside diphosphate to a triphosphate, what is the enzyme?
Why do we need the nucleosides to be triphosphates to incorporate into nucleic acid?
because the polymerases will only use those as starting material
Can you describe the various regulatory roles that AMP and GMP have on their synthesis pathway?
AMP inhibits adenylosuccinate synthetase: This enzyme is responsible for converting IMP to the immediate precursor of AMP, so when AMP levels rise, it directly inhibits its own production.
GMP inhibits IMP dehydrogenase: This enzyme is crucial for converting IMP to the first step in the GMP synthesis pathway, and its inhibition by GMP prevents excessive GMP production.
What are the starting materials to make them?
Bicarbonate, glutamine, ATP, and water
What pyrimidine is made initially?
UMP
What is the role of NDPK? Is it similar to the above use?
phosphorylates UDP to UTP. yes similar use
What does ATCase do?
Aspartate transcarbamoylase (ATCase) is an enzyme that catalyzes a key step in the production of pyrimidine nucleotides.ATCase catalyzes the reaction between L-aspartate and carbamoyl phosphate to form N-carbamyl-L-aspartate
How is it regulated? What activates it and what inhibits it?
Activates: ATP (indicates high energy),high purine levels, and aspartic acid (indicates abundant amino acids)
Inhibits: CTP (indicates pyrimidines are plentiful)
How is this pathway related to the citric acid cycle?
Aspartic acid is readily made from oxaloacetate from the CAC
Why is CTP synthetase regulated by GTP?
Gs and Cs are paired. If we have lots of Gs we need lots of Cs
When nucleotides are catabolized (broken down), what are they disassembled into?
bases and sugars
What is salvage?
reassembly of bases and sugars
Which is more permanent, DNA or RNA?
DNA is more permanent, RNA is more transient
How long does RNA last in the cell?
minutes to hours
What are the steps in taking apart nucleosides?
rna→,nucleoside monophosphate→ nucleoside→ base+sugar
From what nucleoside could we get inosine or hypoxanthine?
adenosine
Why would we get excess uric acid and what disease is caused by this?
It's not water soluble so it's hard to execute, forms crystals in nerves and causes gout
Why is allopurinol a competitive inhibitor of xanthine oxidase and what does it treat?
inhibits the formation of uric acid, is used to treat gout
What is the enzyme HGPRT and what does it do?
recycling the purine bases hypoxanthine and guanine, converting them into nucleotides (inosine monophosphate and guanosine monophosphate respectively) that are essential building blocks for DNA and RNA synthesis
How are deoxyribonucleotides made?
by reduction of nucleoside diphosphates
What does ribonucleotide reductase (RNR) do?
catalyzes the conversion of ribonucleoside diphosphates to deoxyriboside diphosphates
How is RNR regulated? What is the active form?
Needs to be reduced, reduced form is active. ends as oxidized after it catalyzes
What is thioredoxin and how is it activated? What molecule provides the electrons?
reduces RNR to activate it (COFACTOR). gets electrons from NADH
What are the regulatory sites of RNR?
specificity site: controls specificity of enzyme
activity site: controls enzymes activity
How do these sites work?
activity: binding of ATP (stimulatory) or dATP (inhibitory) to the activity site
specificity: influences which substrates will bind to the active site (binding of purines favors pyrimidines in active site)
Which nucleotide triphosphate tends to exist at artificially high levels in the cell? Why?
ATP because cells use it as fuel
Is there any reason to turn off NDPK?
NO
How are thymine nucleotides made?
From Uridine nucleotides. UDP→dUDP→dUTP→dUMP→dTMP→dTDP→dTTP
What is the role of folic acid here?
donates a methyl group for one carbon metabolism
How does the chemotherapeutic drug 5-fluorouracil work?
Suicide inhibitor of thymidylate synthase. If cell can't make DNA and thus divide
How is Folic acid regenerated?
Folic acid is recycled by DHFR
How does the chemotherapeutic drug methotrexate work?
Competitive inhibitor of DHFR, prevents recycling of folic acid and thus prevents cells from dividing
What are synthetic nucleotide analogs used for, medically? How do they work? (More than one strategy.)
treat cancers and viral infections,
fool the polymerase of whatever organism you're putting it into (competitive inhibiting)
act as a chain terminator=> more nucleotides can't be added to the chain
stop replication/kill cell targeting