3 - Nucleotide Chemistry and Metabolism Flashcards
1
Q
Introductory notes in-class
A
- RNA came first - makes sense since it is closer to protein
- Some RNA molecules can act as enzymes even
- They catalyze their own replication - not all RNA molecules do this however
- Originally it was RNA world
- RNA is not stable enough to become the primary code of life
- Coenzymes with nucleotide - e.g. NADH, FADH2, etc.
- Second messengers with nucleotide - e.g. cyclic AMP, etc.
- More focus on Purines over pyrimidines (little clinical importance/implication)
- We need to know passively what they look like ; know which is which at least
- Just know what A, T, G, C, U look like passively; no drawing though
- pYrimidines - thYmine, cYtosine
2
Q
Introductory notes in-class (cont.)
A
- Don’t need to know any unusual bases
- Just be aware that they exist
- Xanthine is a purine
- Caffeine is a purine-like molecule; looks like Adenosine more than anything
- Adenosine can function as neurotransmitter; inhibitory
- Caffeine will competitively inhibit adenosine (which itself is inhibitory), leading to jitters?
- Pregnant women are advised to stay away from caffeine as the embryo may be using analogs instead of the adenosine
3
Q
In-class notes
A
- What does a nucleoside consist of?
- Sugar with a base; ribose and deoxyribose sugars
- What does a nucleotide consist of?
- Nucleoside with a phosphate group (either 1, 2, or 3 phosphates)
- What is 6-amino purine?
- Adenine
- What is 2-amino, 6-oxypurine?
- Guanine
- Where is there a beta-N-glycosidic bond between carbon 1 of a pentose and N9 of a purine or N1 of a pyrimidine?
- Sugar base linkage of a nucleoside
4
Q
The role of nucleotides
A
- Nucleotides are organic units that:
- Serve as the monomers for nucleic acids like RNA and DNA
- Are intermediates in synthesis of carbohydrates, lipids, and proteins
- Components of coenzymes
- Second messengers in signal transduction pathways
- Regulator compounds inhibiting or activating enzymes
5
Q
Nucleotide vs Nucleoside
A
- Nucleotide consists of a nitrogenous base, five carbon sugar (either ribose or 2-deoxyribose), and phosphate group(s)
- Nucleoside consists of the same structure as a nucleotide but lacks the phosphate group(s): nitrogenous base and five carbon sugar (ribose or 2-deoxyribose)
6
Q
Nucleotides
A
- Phosphate groups are responsible for negative charges associated with nucleotides (nucleic acids)
- Ribonucleoside monophosphate, diphosphate, and triphosphate
7
Q
Nucleosides
A
- A nucleoside consists of a nitrogen base added to either ribose or 2‐deoxyribose
- Purine nucleosides end in –osine, while pyrimidine nucleosides end in –idine
- Unless stated otherwise, the sugar is assumed to be ribose
- The purine and pyrimidine numbering systems run in different directions; clockwise vs. counter-clockwise
- Ribose is more common in the cell because it is used in common molecules like ATP and cAMP
8
Q
Nucleotide Terminology
A
- Purine and pyrimidine bases found in RNA and DNA
- Ribonucleotides are nucleotides in which the sugar is ribose
- Deoxyribonucleotides are nucleotides in which the sugar is 2-deoxyribose
9
Q
Chart of Bases and Nucleosides
A
Base: Nucleoside (ribose + base):
uracil uridine
cyotsine cytodine
thymine thymidine
hypoxanthine inosine
adenine adenosine
guanine guanosine
- d-adenosine + Pi –> (deoxy) d-AMP (found in DNA)
- d-thymidine + Pi –> d-TMP
- The above two are deoxyribonucleotides
10
Q
Unusual pyrimidines and purines
A
- Other unusual pyrimidines and purines can occur in DNA and RNA e.g., 5‐ methylcytosine.
- Xanthine derivatives include caffeine, theophylline (found in tea), and theobromine (found in cocoa)
11
Q
What is 6-amino purine?
A
adenine
12
Q
What is 2-amino 6-oxypurine?
A
guanine
13
Q
Synthesis of Nucleotides
A
- Purines: donated carbons and nitrogens are added to a preformed ribose 5‐phosphate.
- Essentially the base is being built as it is being attached to the sugar.
14
Q
Synthesis of nucleotides
A
- Pyrimidines: synthesized before being attached to ribose 5-phosphate
- Essentially the base is assembled and then attached to the sugar.
15
Q
Synthesis of PRPP
A
- Ribose 5-phosphate for nucleic acid synthesis
- 5-phosphoribosyl-1-pyrophosphate is just ribose 5-phosphate with PPi attached to it; that is Pyrophosphate gets the reaction forward
- PRPP synthetase
- Purine ribonucleotide is the product so it is an inhibitor naturally
16
Q
Pentose Phosphate Pathway and Nicotinamide Adenine
Dinucleotide (NADPH)
A
- Glucose 6-phosphate can yield NADPH (for reductive anabolic pathways), ribose 5-phosphate (for nucleic acid biosynthesis), and glyceraldehyde 3-phosphate & F 6-P (for glycolytic pathway)
- Production of NADPH is a irreversible oxidative reaction
17
Q
Purine nucleotide synthesis
A
- The purine nucleotides, AMP and GMP, exert feedback inhibition on PRPP synthetase and PRPP amidotransferase. They also exert feedback inhibition on the reactions leading from IMP to GMP and AMP
- IMP is from hypoxanthine , which is a purine
- Sufonamides are synthetic ; they don’t allow bacteria to make folic acid
- Sulfonamides are structural analogs of para-aminobenzoic acid that competitively inhibit bacterial synthesis of folic acid. Because purine synthesis requires tetrahydrofolate as a coenzyme,
the sulfa drugs slow down this pathway in bacteria. - Methotrexate dirsupt the metabolism of folic acid so further downstream; in the end does the same same, kills the organism;
- These drugs limit the amount of tetrahydrofolate available for use in purine synthesis and, thus, slow down DNA replication in mammalian cells. These
compounds are, therefore, useful in treating rapidly growing cancers, but are also toxic to all dividing cells. - ANTI-FOLATES are the types of the drugs above
- not really used interchangeably
18
Q
Antifolates
A
-
Sulfonamides are structural analogues of PABA that completely inhibit bacterial synthesis of folic acid
- Sulfa drugs stop the pathway that synthesizes folic acid. Kills bacteria, mammals can’t synthesize folic acid so they aren’t affected.
- Methyltrexate (Folic acid analog) limit the amount of tetrahydrofolate available for purine synthesis and thus slow DNA replication in mamalian and bacterial cells.
- PABA Analogs kill bacteria, affect folic acid synthesis of only micro-organisms
- Folic Acid Analogs kill bacteria, affects their metabolism by slowing DNA synthesis of micro-organisms and humans
- Antifolates: sulfonamides and methyltrexate
19
Q
Synthesis of Purine Nucleotides
A
- Conversion of Inosine 5’monophosphate (IMP) to A(adenosine)MP and G(guanosine)MP
- Methods of control:
- Negative feedback, each nucleotide turns off production of itself
- Positive intervention
- Mycophenolic Acid
- Drug is a reversible uncompetitive inhibitor of IMP Dehydrogenase
- Drug deprives rapidly proliferating T and B cells of required nucleic acids, they can’t multiply and this weakens the immune system
- Used to prevent graft infection
- Ultimately you are making nucleotide NOT the bases i.e. AMP and GTP
- Right about of both must be made which is controlled by negative feedback
- Each nucleotide turns off production of itself at critical levels
- Cross-regulation: AMP production require GTP, GTP production requires ATP positive inhibition/intervention
20
Q
De Novo Sythesis of Purines
A
21
Q
Regulation of purine synthesis
A
My note says Dr. Marvit told us not to memorize this figure
22
Q
Folic Acid
A
- Folic acid deficiency is the most common vitamin deficiency today in the USA.
- Folic acid supplementation before conception and during the first trimester can help reduce the risk of neural tube defects (including spina bifida)
- Tetrahydrofolate and its derivatives function in the carriage and transfer of one carbon units in the biosynthesis of certain amino acids, purine nucleotides, and dTMP
- Spina bifida and other neural tube defects affect about 4,000 pregnancies in the USA each year
- Methotrexate is a folate analogue that competitively inhibits dihydrofolate reductase
- Methotrexate is used for the treatment of psioriasis, rheumatoid arthritis, and certain types of cancer
- What is the “common denominator” of these diseases? They involve cell divisions which require purines.
- Psioriasis, rheumatoid arthritis are all auto-immune diseases; all go through B and T rapid division and use a lot of purines
23
Q
Salvage Pathways for Purines
A
- Purines that result from the normal turnover of cellular nucleic acids, or that are obtained from the diet and not degraded can be reconverted into nucleoside phosphates.
- The salvage pathways represent the only source of purine nucleotides for tissues that are not capable of synthesizing purines de novo
- While most tissues can synthesize purines de novo, this process is most active in the liver. The liver subsequently exports the products to other portions of the body.
- DNA is NOT an essential nutrient nor is RNA
- Our body will make it from destroying the old ones; recycling i.e. salvage pathways
- HGPRT enzyme doesn’t function –> x-linked recessive inherited disorder
- PRPP will build up as a result and its build up induce DE NOVO synthesis ; since GMP or AMP levels are down; excessive de novo purine synthesis
- When purines are degraded, they are made into uric acid NOT Urea
- Uric acid is a relatively powerful antioxidant; breakdown product functions as anti-oxidant
however, too much uric acid is BAD - Lesch-Nyhan Disease
24
Q
Digestion of dietary
nucleic acids
A
25
DNA Molecule
Per Dr. Marvit: just know that there are 3,5 phosphodiester bonds there ; no details
26
Degradation of Purine Nucleotides
* Degradation of dietary nucleic acids occurs in the small intestine, where a family of pancreatic enzymes hydrolyzes the nucleotides to nucleosides and free bases. Inside cells, purine nucleotides are sequentially degraded by specific enzymes, with uric acid as the end product of this pathway.
* *Adenosine Deaminase (ADA) Deficiency*
* Causes severe combined *immuno deficiency (SCID)*, involves both T and B cell disfunction
* Extremely large *buildups of dATP*
* Gout
* Treatment with allopurinal inhibits xanthene oxidase, results in accumulation of hypoxanthine and xanthene which are more soluble than uric acid
27
Gout
* Gout is uric acid crystals in the joint, typically of the big toe
* Why big toe? Because it is peripheral and at a colder temperature than rest of body. Crystals precipitate in decreased temp
* Avoid alcohol, is a diuretic, lose water, uric acid is less soluble and can crystalize
* Hyperuricemia is coniditon of too much uric acid in the blood. Humans typically have close to the maximium amount of uric acid in their blood because it is an antioxidant.
* Primary Gout is an increase in uric acid because you have an increase in purines
* Seconday Gout is much more common and is caused by other diseases
* Secondary hyperurocemia due to high cell proliferation --\> gout (since it requires a lot of purines)
* Typically associated with cancer, anemia, sporiasis
* Glycogen diseases lead to problems with sugars (ribose 5-phosphate), which leads to problems with nucleic acid synthesis
* Bottom line, purines can NOT be stored in body and eventually lead to gout
* *Allopurinol* treatment is the enzyme inhibitor of xanthene oxidase. This stops the degradation of xanthene and hypoxanthene to uric acid.
* Purines can't be stored so they must be degraded
28
Gout
* Gout can be caused by either the overproduction or
underexcretion of uric acid, the end result being the deposition of monosodium urate crystals in the joints and a subsequent inflammatory response
* While the overproduction of uric acid does not account for the majority of cases of gout, this overproduction can be caused by any of the following:
* Mutation in the X‐linked PRPP synthetase gene, resulting in an increased Vmax, a lower Km, or a decreased response to allosteric inhibitors
* Lesch‐Nyhan Disease: a rare, X‐linked, mutation of HGPRT which leads to the inability to salvage hypoxanthine or guanine. The resultant increase in PRPP and decrease in GMP can lead to an increased de novo purine synthesis and ultimately an increase in levels of uric acid
* Lesch‐Nyhan is often associated with self mutilation. The reasons for this self mutilation have not been entirely elucidated
29
Arthrocentesis
* Joint aspiration, a procedure whereby a sterile needle and syringe are used to drain fluid from a joint. Can be used to diagnose gout
* Monosodium urate crystals in aspirated synovial fluid
30
Ribonucleotide Reductase
* Ribonucleotide reductase regulates the balance of the 4
deoxyribonucleotides required for the synthesis of DNA
* Hydroxyurea is an inhibitor of ribonucleotide reductase.
Hydroxyurea is used in the treatment of certain cancers (via stopping production of DNA), myeeloproliferative disorders, and also of sickle cell anemia (independent of ribunucleotide reductase)
* Uric acid is bigger than urea
* Sickle cell increase hemoglobin F, which has *nothing* to do with what's shown here
* In the figure, it is easier to remove water than just trying to remove O; NADPH is the source of H
31
Adenosine deaminase (ADA) deficiency
* Lymphocytes express the highest activity of adenosine deaminase. A deficiency in this enzyme can inhibit ribonucleotide reductase, resulting in B‐, T‐, and natural killer-cell deficiency. This is known as severe combined immunodeficiency disease (SCID). About 1 out of every 7 cases of SCID result from ADA deficiency
* A deficiency of adenosine deaminase can lead to a buildup of adenosine which can be converted into dATP. This dATP will inhibit ribonucleotide reductase, subsequently preventing the synthesis of all deoxyribonucleotides.
* Some cells more dependent than others on ADA so they are affected unequally
* First disease to be benefiting from genetic therapy
* Very rare disease 1/10 of freq. of PKU
* Adenosine deaminase has other substrates that is required for proper function , which is dAMP, ultimately metabolized into dATP in normal pathway function.
* If ADA is missing or dysfunctional, dAMP is built up
* Bottom line, dATP is built up too much and no more DNA can be properly built for synthesis
* Adenosine itself doesn't inhibit the process; normal ATP, AMP, cAMP doesn't affect
* ITS ONLY THE dATP that is the culprit
32
Pyrimidine Synthesis
Unlike the synthesis of the purine ring, in which the ring is
constructed on a preexisting ribose 5‐phosphate, the pyrimidine ring is synthesized before being attached to ribose 5‐phosphate, which is donated by PRPP.
33
Antimetabolites
* Antimetabolites inhibit a specific metabolite
* The include:
* Purine analogues (competitive inhibition)
* Pyrimidine analogues
* Anti‐folates (i.e. sulfa drugs, methotroxylate; affect DNA transcription & replication also)
* Nucleoside analogues
* They disrupt DNA replication, transcription, or purine synthesis
* These anaolgues can be used in:
* Cancer chemotherapy (depriving of DNA)
* Immuno suppression (for transplant)
* Viral inhibition
* Aciclovir is used as an anti-herpetic agent
34
Summary
35
Clinical questions
1. How might glycogen storage disease lead to gout?
2. Why do the sodium urate crystals tend to form in the big toe? - relatively coldness
3. How might malignancies and/or chemotherapy cause gout? - buildup of purines, breakdown of purines; urine
4. How might the ingestion of alcohol aggravate gout? - losing water
