Purines and pyrimidine

Purines

Xanthinine, hypoxanthinine, adenine, guanine

Pyrimidines

Cytosine, Uracil, Thymidine, Orotic Acid

Difference of structure between Purines and Pyrimidines?

Purines have a fused 5 membered and 6 memebred ring

Pyrimidines have one 6 membered ring

Which nucleotides are not found in DNA?

A. Hypoxanthinine

B. Thymine

C. Cytosine

D. Uracil

E. Guanine

F. Xanthinine

A, D, F

What is the structure called when a sugar is added to a nitrogen base?

Nucleoside

Which of the following is not a nucleoside?Adenosine

Guanosine

Inosine

Hypoxanthine

Hypoxanthine

Which of the following is not a nucleoside?

Uridine

Thymidine

Adenosine

Thymine

Cytidine

Thymine

Which is the nucleoside which is the purine?

Thymine vs thymidine?

Purine 2

Nucleoside 1

Name 4 Purine Nucleotides

AMP, GMP, dGMP, aATP

What is it called when you add one or more phosphates to a ribose and a nitrogen base?

Nucleotide

Nucleotide vs nucleoside

Guanine vs guanosine

NT NS

What are the products of degrades purines?

Urate and uric acid

What are the products of degraded pyrimidines?

NH4 + CO2 + RCOOH

Where is the enzyme xanthine oxidase present? What reaction does it catalyze? How is it inhibited?

Bonus point, relate to med chem!

Present in the liver and intestine, catalyzes reaction of hypoxanthine into xanthine than xanthine into urate, inhibited by allopurinol at the active site as a competitive inhibitor, inhibited by uloric at the allosteric site as a non competitve inhibitor.

Xanthine oxidase works on aldehydes to make carboxylic acids

What are the four essential roles of PRPP?

De novo purines

De novo pyrimidines

Salvage pathway of purines

NAD AND NADP formation

What are the precursors to PRPP and why is this important?

Sugar molecules must be activated in order tp catalyze a reaction. ATP transfers a phosphate group to activate the sugar into a pyrophosphotate which is essential for activating sugars and carbohydrates also in the case of glycosylation

Describe the enzyme kinetics of glutamyl PRPP aminotransferase for purine synthesis

This shows a sigmoidal curve, the concentration of PRPP MUST be above the KM for the substrate or else the reaction will not occur. If enough PRPP is not available de novo synthesis does not occur. High concentrations of PRPP can overcome the inhibition of the nucleotides. The flux can increase when an increase in substrate occurs.

Comparisons in purine and pyrimidine synthesis

And contrasts?

Both require synthesis of PRPP, activated by high energy phosphate bonds in order to activate the ribose 5 phosphate into an activated sugar, inhibited by AMP

Contrast: in de novo synthesis, NO OR INSUFFICIENT HYPOXANTHINE AVAILABLE SO NO SALVAGE OCCURS! In return concentrations of PRPP rises activating GPRPPAT

In salvage pathway hypoxanthine or guanine must be available

What do purines require during synthesis from PRPP to IMP? (6)

N10 formyl THF, glycine, aspartate , ATP, glutamine, CO2

What is the nucleobase of IMP?

Hypoxanthine

In the synthesis of purine nucleotides, how does the body control the synthesis of AMP vs GMP?

when AMP synthesis is high, 2 phosphates are added to form ATP and this inhibits the synthesis of AMP and activates the synthesis of GMP, GTP is required for AMP synthesis and ATP is required for GMP synthesis

What is the rebate limiting enzyme of purine nucleotide synthesis? How is it activated and how is it inhibited?

Glutamine PRPP aminotramsferase requires high levels of PRPP to be activated and glutamine and is inhibited by high amounts of AMP and GMP which signals the cell it has synthesized enough nucleotides

What reaction does Adenosine deaminase catalyze?

The breakdown of adenosine into ionisine

How does methotrexate inhibit purine biosynthesis? What endogenous substrate inhibits the same reaction?

Methotrexate depletes the cell of THF stores therefore no THF is provided to the cell that is required for the synthesis of PRPP to IMP, no IMP is made therefore the cell is deficent in purines.

High levels of ADP also inhibit this reaction because the de novo synthesis requires ATP

What substrate determines the de novo or salvage of purines?

PRPP

What substrates does Purine Nucleotide Phosphorylase use for the synthesis of hypoxanthine and ribose 5 phosphate?

Xanthonine, Ionisine, Guanosine

What substrates build up in an ADA deficiency?

AMP ATP dATP, ADP dADP, dATP inhibits the synthesis of all deoxyribonucleotides by inhibiting ribonucleotide reductase

What must be recycled to inhibit the overproduction of urate in the body? (2)

Hypoxanthine and guanine

What reaction does HGPRT catalyze? What happens if someone is deficient?

HGPRT catalyzes the reaction of hypoxanthine and PRPP into IMP for the salvage pathway of purines, the preferred mechanism of purine synthesis in the body to decrease urate production. Deficiency leads to LN syndrome and build up of hypoxanthine and deficiency of guanine occurs, bout results due to inability to salvage purines

How does degradation of purines occur, specify location and enzymes

Cell breaks down AMP to IMP via ADA than IMP to hypoxanthine via PNP, hypoxanthine travels in the blood where it is taken to the liver. The liver contains xanthine oxidase to take hypoxanthine to xanthine and again to urate. The liver pumps urate into the blood where it is taken to the urine and excreted as uric acid

Under what circumstances would someone be more likely to degrade purines?

Lack of phosphate like fructose or lactose deficiency, lack of fuel, starvation, alcoholics, tumors lysis syndrome, hypoxia

What reaction does PNP catabolize?

The breakdown of nucleoSides into nucleobase and ribose sugar. Nucleobase snare released into the blood for salvage

What is a suicide substrate for the synthesis of urate?

Oxypurinol, kills xanthine dehydrogenase

True or False, all urate ends up in the glomerular filtrate?

True

How much urate is excreted by the kidney?

7-10%

What is a body builder at risk of and why? How can we help him?

Since he is building up in lactate during his intense workouts he is at risk for hyperuricemia because the build up of organic acids like lactate, betahydroxybuterate, and a acetoacetate will compete with the organic acid transporters for excretion into the kidneys. The amount of urate in his blood will be more than 8 mg/dL but the amount of urate in his urine will be less than 600. We can help him by giving him probenecid if he doesn't have any risks of kidney stones or kidney failure. This will activate the organic acid transporters to aid in the excretion of the organic acids

True or false, the production of urate from xanthine is reversible

False, irreversible that's why salvage is required

What substrates does HGPRT use for salvage and what are the products?

Hypoxanthine into IMP and guanine into GMP

True or false, the salvage pathway undergoes tight regulation

True, the salvage pathway undergoes regulation requiring hypoxanthine or guanine as substrates as well as PRPP, low ADP and high phosphate

What feedback regulates HGPRT?

Nothing! Trick question!

What enzymes shows coopertivity for PRPP and requires a high concentration for activation?

De novo synthesis of purines by glutamine PRPP aminotramsferase

Who is at highest risk for hyperuricemia? Who has the lowest plasma urate?h

Men and post menopausal women. Lowest children

How does someone develop gout?

Prolonged hyperuricemia builds up and forms ionizable sodium salt deposits and lay in the synovial joint fluid causing precipitates. Neutrophils engulf the crystals and cause inflammation. Treated with ibuprofen and indomethacin

Who is at highest risk for overproduction of urate?

Obesity, psoriasis, cytolysis therapy, hemolytic disease, alcohol abuse. Higher levels of uric acid in the urine around 600mg

Treatment for someone who aquires hyperuricemia from cytolytic therapy?

Allopurinol in order to excrete more hypoxanthine, inhibits xanthine dehydrogenase. Or rasbiuricase which is used to make urate into Allatoin to allow easier excretion, much more water soluble. Side effect is its very IMMUNOGENIC

Under what conditions would it be possible that someone has a normal production of urate but still has hyperuricemia?

This would be if someone has lead poisoning, organic acid production, diuretic therapy, low dose aspirin and kidney failure.

Rx: probenecid except for this susceptible to kidney stones

Hyperuricemia with alcohol consumption?

The break down of alcohol requires a lot of ATP leaving the cell with high levels of AMP to activate the degradation of purines at a high rate. The cells are depleted of their NAD+ stores shifting the metabolism to produce lactate therefore leading to decreased excretion of uric acid

How is de novo synthesis facilitated in hyperuricemia for those with a G6Pase deficiency

A build up in the intermediates of glycolysis leads to higher levels of ribose 5 phosphate leading to increased synthesis of PRPP with independently activates the de novo synthesis pathway. Simultaneously lactic acidosis occurs due to low energy production and inability to utilize glycogen

How do chronic hemolytic anemias contribute to hyperuricemia?

Purine de novo synthesis overproduction because of high purine turnover as well as increased degradation due to hemolytic disease

Ex. G6PD def, sickle cell, PK def

Liver cells with a very low concentration of cytosolic free phosphate have a high rate of

A. de novo synthesis of purine nucleotides.

B. oxidation of xanthine to urate.

C. salvage of hypoxanthine.

D. synthesis of OMP (orotidine monophosphate).

E. synthesis of PRPP (phosphoribosyl pyrophosphate).

A. Wrong for this to occur the cell requires high levels of phosphate, low levels of ADP,low levels of hypoxanthine and high concentrations of PRPP

B. CORRECT!

C. False, this would be true if the cell had PRPP and enough energy to synthesize PRPP, low ADP

D. False this is pyrimidine synthesis with PRPP and oroate as substrates

E. False, this requires high levels of free phosphate and low levels of ADP

Most cells can synthesize purine nucleotides de novo. This pathway has a high rate of flux.

True or false

False, this has a very low rate of flux because it requires a state of high energy and high concentration of metabolites. Glycine, glutamine, THF, CO2 aspartate, ATP

True or false. Purines are built step-wise on DNA bases like guanine and adenine

False, they are built off of ribose 5 phosphate from PRPP

True or false.

Cells generally re-use most of the hypoxanthine in the blood to replenish their own nucleotide stores. In normal individuals, the flux in the salvage pathway is much larger than in the de novo synthesis pathway.

True

True or false

Urate production is normal with uric acid levels in the urine of <600 mg in patients with excessive degradation of purines (cytolytic therapy, psoriasis, obesity, alcoholism, chronic hemolysis, etc.)

False, urate production is high with urine concentration of > 600

True or false

TMP can be synthesized de novo.

False, UMP is synthesized de novo and is used to synthesize CTP from UTP and dTTP by dUMP

True or false

Hydroxyurea and gemcitabine are chemotherapeutic drugs that inhibit ribonucleotide reductase which uses UDP CDP TDP and ADP as substrates

False, this enzyme does not use TDP, everything else is correct

True or false

Synthesis of purine nucleotides is controlled mostly at the level of PRPP synthetase, CAP

synthetase II, and ribonucleotide reductase.

False, this is pyrimidines

True or false

Carbohydrates are activated for the synthesis of polysaccharides like glycogen and protein/lipid glycosylation through conjugation to ADP.

False, conjugated to UDP

Membrane components such as phosphydilcholine require what for their activation?

They require CDP conjugation to Choline to be activated

What is the importance of glycoslated glucose by UDP?

When glucose in conjugated by UDP this provides energy by a pyrophosphate for the synthesis of glycogen. It can also be used in enzymatic reactions to serve as a high energy substrate for producing components like plasma membrane and lipids and proteins

What provides the pyrimidines ring during de novo synthesis?

Orotic acid

What are the pyrimidine nucleobase matched with their nucleoside?

Nucleobase: orotic acid, thymine, uracil, cytosine

Nucleoside: oritidine, thymidine, uridine, cytidine

What is the role of the liver in pyrimidine synthesis?

Pyrimidine synthesis takes place in thr

True or false

PRPP is increased in concentration in an ADA deficency

False

What are two agents used to inhibit the synthesis of dTMP?

5-florouracil and methotrexate

What is a suicide substrate for thymidine synthetase?

5-F-dUMP

What is the mechanism of 5-F-UTP?

It is added into RNA and misincorporated to hault RNA synthesis

Is ATP required for the salvage pathway of pyrimidines?

Yes ATP is required in order to transfer a phosphate group in order to keep the uridine inside of the cell in the form of UMP!

What quenches the essential tyrosyl radical of ribonucleotide reductase?

Hydroxyurea

What is a suicide inhibitor of RNR?

Gemticinbine

When the cell is high in AMP, what is the concentration of purine nucleotides?

High enough and we do not need to synthesize them or the cell is low in energy

When a cell has low concentrations of AMP what is the energy state and concentration of nucleotides?

Enough ATP and well energized and need more nucleotides for synthesis of DNA or energy.

True or false:

There is an independent mechanism to activate the synthesis of purine pyrimidine nucleotides

True, this is the function of PRPP

Where is the flux of pyrimidine synthesis the largest? Where does it normally occur?

Largest in replicating cells, occurs in liver and replicating cells

Is there a feedback inhibition of pyrimidine syntheses?

Yes, by UTP on CPS2

How do you make cytidine from uridine?

Synthesis of uridine to UMP than diphosphorylate it to make UTP than use glutamine to replace the ketone of UTP with an amine group to make CTP which is dephosphorylated to make CDP

What is the molecule that acts as an alkylation agent of RNR?

Gentimidne