Lecture 30 Flashcards
What is the important precursor for all nucleotides?
PRPP
Compare purines and pyrimidines in terms of their PRPP involvement, pathway, and the location in the cell where they occur.
Purines:
Form nitrogenous base on PRPP
Branched pathway
Cytoplasmic
Pyrimidine:
form nitrogenous bases independent of PRPP
unidirectional pathway
Cytoplasmic
Compare purines and pyrimidines in terms of their precursors used, and Regulation method
Purines: Precursors: -NH3 from Gln Gly, Asp N10-formyl-THF HCO3-
Regulation:
Feedback inhibition by purines
Pyrimidines: Precursors: -NH3 from Gln Asp HCO3-
Regulation: (both are allosteric)
Pyr inhibits it (C)
Pur activates it (A/G)
Briefly describe what CAD is and the enzymes it uses to synthesize ribonucleotides.
CAD: multifunctional eukaryotic protein that performs the beginning steps of pyrimidine synthesis (contains the following enzymes)
Carbamoyl phosphate synthetase II (CPSII)
Aspartate transcarbamoylase (ATCase)
Dihydroorotase
(CAD)
Which of the 3 enzymes that compose CAD features “a channel”?
CPSII (Carbamoyl phosphate synthetase) has a channel
State what activates/inhibits ATCase (aspartate transcarbamoylase). For dihydroorotase, state its function and where in the cell it is located.
ATCase is activated by ATP and inhibited by CTP
Dihydroorotase acts as a reverse hydrolase to close a ring (makes dihydroorotate from Carbamoyl Aspartate) it it does this in the mito
State the 2 functions of UMP synthetase
removes PPi when orotate is added to PPRP
Decarboxylates orotate to form uracil
Megaloblastic anemia occurs when cells fail to _____. What are 2 common causes of this?
divide
B9 and/or B12 deficiencies
Deficiency of what enzyme is known to cause Hereditary Orotic Aciduria? State 2 symptoms of this condition
deficiency of UMP synthetase
- Excessive orotate in the urine
- Megaloblastic anemia that fails to respond to B9/B12 treatments
Compare the specificities of Nucleoside monophosphate kinases and Nucleoside Diphosphate kinases during Pyrimidine synthesis
Nucleoside monophosphate kinases are specific to each NMP
Nucleoside Diphosphate kinase has a broad specificity
Compare what activates and inhibits CTP synthase and ATCase. Which of these activating/inhibiting molecules is the only nucleotide to be synthesized directly as a triphosphate?
CTP Synthase:
GTP activates and CTP inhibits
ATCase:
ATP activates and CTP inhibits
CTP is the only nucleotide to be synthesized directly as a triphosphate
From PRPP, describe the 10 steps that form the IMP molecule that purines are made from
- An NH3 group replaces the PPi group on PRPP
- Glycine is added
- A Formyl group, from N10-formyl THF is added to complete the ring
- An NH3 group from Gln is added (to start the 2nd ring)
- The 2nd ring is closed
- 2 CO2 molecules, both from HCO3 are added. First to Gln (N) and then to Gly (alpha-C)
- Asp is added to the Carboxyl
- Fumarate is released
- A 2nd Formyl group addition occurs, also from N10-formyl THF (preparing to close the ring)
- The 6 membered ring is closed to form IMP (Hypoxanthine?)
When synthesizing purines, describe the 3 enzymes that carry out steps 1,4, and 8 and then describe the 3 enzymes that conduct all the other steps. What is the final product of all 10 steps of purine synthesis?
3 individual enzymes carry out steps 1, 4, and 8
3 multifunctional enzymes carry out all of the other steps
IMP is the final product
Since both AMP and GMP are synthesized from IMP, compare their formation in terms of what they use for energy, what inhibits their formation, and a basic description of what functional groups are altered
AMP: uses GTP for energy and is inhibited by AMP
NH3 from Asp replaces it’s carbonyl and Fumarate is released
GMP: uses ATP for energy and is inhibited by GMP
Redox with H2O creates a 2nd carbonyl that is quickly replaced by an NH3 group from Gln
Converting ribose to deoxyribose requires a ______ reaction. Where do the electrons necessary for this reaction come from?
reduction
The electrons come from NADPH
Explain how Ribonucleotide reductase acts on NDPs and NTPs. How does this enzyme pass electrons and what stabilizes this process?
Ribonucleotide reductase acts on ALL NDPs to form dNDPs
Passes electrons via the creation of free radicals which are stabilized by 2 Fe Ions
In terms of the regulation process of Ribonucleotide reductase, which allosteric site acts as the “on off switch” and which allosteric site acts as the “volume dial”?
Activity site = on/off switch
Specificity site = volume dial
(these are both Allosteric sites, NOT the “active site” of the enzyme) (the name activity site is misleading)
When regulating ribonucleotide reductase, explain the effects ATP and dATP have on it’s activity by way of interacting with the activity site.
ATP stimulates ribonucleotide reductase activity (on switch)
dATP inhibits ribonucleotide reductase activity by altering subunit contacts (off switch)
When regulating ribonucleotide reductase, explain the effects dATP, dTTP, and dGTP have on it’s activity by way of interacting with the specificity site.
dATP: pyrimidine formation is preferred
dTTP: GDP is preferred (pyrimidine formation inhibited)
dGTP: ADP is preferred (pyrimidine formation inhibited)
What does Nucleoside diphosphate kinase do to dNDPs?
makes them into dNTPs
Describe the 2 different reactions that can yield dUMP from either dUTP or dCMP.
Removal of PPi from dUTP yields dUMP
Deamination of dCMP yields dUMP
Compare the functions of endonucleases and exonucleases
Endonucleases: cut in the middle of a nucleic acid strands
Exonucleases: chew from the end of a nucleic acid strand
Pyrimidines use _____ and _____ for salvage in 2 steps. State the molecule that the activity of these 2 blanks will yield.
Phosphorylases
Kinases
Phosphorylases make nucleoSIDES and Kinases make nucleoTIDES
Describe what is dangerous about viral thymidine kinase, compared to human thymidine kinase. What substance can help prevent the danger presented by viral thymidine kinase?
Viral Thymidine Kinase is less discriminating than human Thymidine Kinase, so it will accept purines as well as T’s
Acyclovir will selectively bind to viral thymidine kinase (doesn’t bind to human thymidine kinase) and terminate it’s synthesis
What enzyme do purines use to salvage? explain what is yielded after salvaging Adenine, Guanine, and Hypoxanthine
Purines us phosphoribosyltransferase to salvage
Adenine + PRPP = Adenylate + PPi
Guanine + PRPP = Guanylate + PPi
Hypoxanthine + PRPP = inosinate + PPi
state the final products of both purine and pyrimidine catabolism
Purine catabolism yields Uric Acid
Pyrimidine catabolism yields Beta-Ureidopropionase acid
What is the mechanism of gout?
Chronically elevated urate levels in the blood causing uric acid crystals to form in the joints
Causes inflammation, arthritis, and joint degradation
What is Lesch-Nyhan Syndrome a deficiency of?
HPRT
SCID (severe immunodeficiency disease) is actually a deficiency of what? describe the mechanism of this disease
Adenosine deaminase
without it, dAMP accumulates and is converted to dATP by salvage pathway enzymes
dATP then inhibits ribonucleotide reductase, which means DNA is no longer able to be synthesized
Which Deoxyribonucleotide is the target of many anti cancer chemotherapy drugs? State the 2 enzymes that are common targets of these drugs
dTTP
- Thymidylate synthase
- Dihydrofolate reductase
State the 9 steps of de novo pyrimidine synthesis
- CPSII adds an NH2 from Gln to Bicarbonate + ATP to make Carbamoyl Phosphate
- ATCase adds Asp
- Dihydroorotase closes the ring in a dehydration step
- Dihydroorotate dehydrogenase (in the mito) forms orotate (from dihydroorotate)
- UMP Synthase removes PPi as orotate adds to PRPP
- UMP Synthase conducts a Decarboxylation to make uracil from orotate
- UMP becomes UDP via Nucleoside monophosphate Kinase
- UDP becomes UTP via Nucleoside Diphosphate Kinase
- CTP Synthase makes CTP from UTP
