Lecture 9 - Amino Acid And Nucleotide Breakdown And Synthesis

Question 1

What are the 3 circumstances under which amino acid degradation would occur?

Answer 1

1) eat a protein rich meal and the bodily demand for AAs is less than was ingested
2) starvation or uncontrolled diabetes
3) normal cellular turnover

Question 2

How is cellular turnover of proteins divided further?

Answer 2

1) housekeeping - break down proteins that have not folded properly and accumulation would be harmful
2) regulatory - to control cellular metabolism

Question 3

What does the rate of protein turnover tell about the protein?

Answer 3

Proteins that are rapidly degraded represent important metabolic control points

Question 4

How is protein degraded in the tissue? (2 ways)

Answer 4

Lysosomal degradation

Proteasome degradation via ubiquitination

Question 5

How are proteins selected for lysosome degradation?

Answer 5

Chaperone mediated autophagy (CMA)

Question 6

When will the cell use CMA to degrade protein?

Answer 6

Under starvation conditions when proteins are needed for energy

Question 7

How are proteins selected for CMA?

Answer 7

They are tagged for degradation via KFERQ sequence and chaperones recognize this sequence on proteins in the cytosol and bind to the protein and transport it to the lysosome for degradation

Question 8

Describe the process of CMA and the role of LAMP-2A.

Answer 8

Cytosolic chaperone recognizes KFERQ sequence and binds to protein. Chaperone brings protein to lysosome surface to LAMP-2A receptors at the surface of the lysosome. LAMP-2A monomers aggregate to form multimer complex which then move the protein into the lysosome for degradation.

Question 9

How is CMA regulated?

Answer 9

LAMP-2A levels control CMA activity, upregulation of LAMP-2A leads to increased CMA (occurs during starvation) downregulation results in decreased CMA activity

Question 10

Under what conditions will the cell utilize the proteasome for degrading proteins?

Answer 10

Housekeeping and regulatory situations

Question 11

Describe the process of degradation by the proteasome

Answer 11

The protein must first be ubiquitinated by 4 ubiquitin molecules. Then that protein binds to the 19S cap of the proteasome. ATPases in the 19S cap will hydrolysis ATP to unfold the protein. The protein is then passes to the 20S core where it is degraded by enzymes into AAs. The ubiquitin is recycled.

Question 12

What determines how long a protein will exist in the cell? (2)

Answer 12

The N-end rule: the AA that occupies the N terminal position plays a role in the stability of the protein

PEST proteins: proteins that have segments rich in proline, glutamate, serine or threonine may provide sites for phosphorylation which promote ubiquitination

Question 13

When an individual amino acid is degraded, what are the 2 major products of that degradation?

Answer 13

1) Ammonia (NH3) —> becomes urea
2) Carbon skeleton —> can be oxidized for energy to CO2 + H2O, can be used to make glucose via gluconeogenesis, can be used to make acetyl CoA or can be used to make ketone bodies

Question 14

Where is urea synthesized?

Answer 14

The liver and kidney to a small extent

Question 15

What are the 2 means that nitrogen groups enter the urea cycle to produce urea?

Answer 15

As carbamoyl phosphate or as aspartate

Question 16

Which parts of the urea cycle occur in the mitochondria? In cytosol?

Answer 16

Glutamate dehydrogenase
Aspartate amino transferase
Carbamoyl phosphate synthetase1

Remainder in cytosol

Question 17

How does non hepatic tissue excrete nitrogen?

Answer 17

Process of transdeamination occurs in the tissue per usual, but the L-glutamate formed is transformed into L-glutamine so it can travel in the blood to the liver (glutamate is excitatory NT, can’t travel in blood) where it is converted back to L-glutamate

Question 18

When does the glucose alanine cycle occur?

Answer 18

When a muscle tissue is actively breaking down itself (it’s own protein)

Question 19

Describe the glucose alanine cycle

Answer 19

Glucose in muscle is broken down to pyruvate via glycolysis. The Nitrogen containing group from break down of an AA is transferred to pyruvate via transamination to form alanine. Alanine then travels to liver via blood where it is converted back to pyruvate and in the process it’s attached Nitrogen group is excreted as urea. The pyruvate can then be used to regenerate glucose which is transported back to the muscle.

Question 20

How is the urea cycle regulated in the short term?

Answer 20

Allosteric activation of carbamoyl phosphate synthetase by N-acetylglutamate (synthesized from glutamate)

+ AA break down —> + [glutamate] as a result of + transamination —> + [N-acetylglutamate] —> activates urea cycle

Question 21

How is the urea cycle regulated long term?

Answer 21

Prolonged starvation and high protein diets lead to increased need for the urea cycle enzymes to function so there is an increased rate of synthesis of these enzymes

Question 22

In terms of the carbon skeleton of amino acids, how are these classified?

Answer 22

Either ketogenic (forms ketone bodies) or glucogenic (forms glucose), can be classified as both

Question 23

With regards to amino acid biosynthesis, where do the precursors of the carbon skeleton come from? Where to the amino groups come from?

Answer 23

Derived from intermediates of glycolysis, the CAC and PPP

Glutamate and glutamine

Question 24

What is an essential AA? Non essential AA? What makes an AA essential?

Answer 24

Essential = body cannot synthesize naturally, must be obtained from food

Nonessential = body can synthesize naturally

Lack of enzymes to produce, inability to produce the needed amount

Question 25

What are the 2 ways nucleotides can be made?

Answer 25

De novo synthesis (from scratch)

Salvage pathway (recycle nucleotides)

Question 26

What is the role of formate in nucleotide synthesis?

Answer 26

Formate donates single carbons

Derived from Tetrahydrofolate, which is driven from the vitamin folic acid

Question 27

Describe the synthesis of purines de novo

Answer 27

Ribose 5 phosphate from pentose phosphate pathway is activated using ATP to form PRPP which is used to synthesize nucleotides or certain AAs. Then the nitrogen double ring is built on top of this sugar to form IMP. IMP can then be made into AMP or GMP, which can be phosphorylated by nucleoside monophosphate kinases and nucleoside diphosphate kinases to form ADP ATP and GDP GTP.

Question 28

How is purine synthesis regulated?

Answer 28

1) synthesis of PRPP is inhibited by ADP
2) AMP, GMP and IMP inhibit the 1st committed step in their synthesis which is catalyzed by glutamine PRPP amidotransferase
3) AMP and GMP independently regulate their own production
4) increased levels of PRPP activate production of AMP and GMP

Question 29

Which enzyme is the 1st committed step of purine synthesis?

Answer 29

Glutamine PRPP amidotransferase

Question 30

Describe de novo synthesis of pyrimidines

Answer 30

NH4+ combines with HCO3- and ATP is hydrolyzed to form carbamoyl phosphate via the enzyme carbamoyl phosphate synthetase II. Then, CP combines with Aspartate to form N-carbamoylaspartate. Several more steps lead to the production of an intermediate derived from N-carbamoyl aspartate that combines with PRPP to form UMP. ATP can be used to form UDP and then UTP, which can be used to form CTP which can be used to form CMP.

TTP must be formed via a different route

Question 31

How is pyrimidine synthesis regulated? (2)

Answer 31

1) High [PRPP] stimulates (feed forward) the production of N-carbamoyl aspartate
2) Increased [UDP] and [UTP] inhibit carbamoyl phosphate synthetase II

Question 32

What enzyme catalyzes the conversion of a ribonucleoside into a deoxyribonucleoside?

Answer 32

Ribonucleotide reductase

Question 33

Describe how hydroxyurea inhibits ribonucleotide reductase.

Answer 33

During the enzyme’s mechanism of action, a Tyr residue on the enzyme forms a tyrosyl radical which promotes the formation of a radical in the enzyme’s active site.

Hydroxyurea scavenges the radical at the active site of the enzyme preventing the biosynthesis of deoxyribonucleotides.

Question 34

What medical application utilizes hydroxyurea?

Answer 34

Cancer treatment

Question 35

How is thymine synthesized?

Answer 35

dUMP is first formed via mechanisms discussed earlier. Then, thymidylate synthase uses THF as a coenzyme to form dTMP which can then be phosphorylated to form dTDP and dTTP as needed.

Question 36

How can drugs be used to target the synthesis of dTMP for cancer treatment?

Answer 36

Anti folate drugs can be used since folate is necessary for synthesis of dTMP

Question 37

What are the 2 enzymes in the folate cycle that can be inhibited for cancer treatment?

Answer 37

Thymidylate synthase can be inhibited by fluorouracil, which is a suicide inhibitor (covalently binds to active site and destroys functionality of enzyme)

Dihydrofolate reductase can be inhibited by competitive inhibitors that resemble substrate (7,8 dihydrofolate)

Question 38

What are the 2 sources of Nucleic acids that would need to be degraded?

Answer 38

1) Nucleic acids from diet

2) Nucleic acid turnover (recycling)

Question 39

Summarize purine catabolism.

Answer 39

Nucleotides are broken down into nucleosides, which are all then broken down into Xanthine by Xanthine oxidase (XO). XO then converts Xanthine to Uric acid which is excreted.

Question 40

What is gout? How is it caused?

Answer 40

Gout is the accumulation of uric acid crystals in the joints. It can be due to impaired excretion of uric acid or over production of uric acid. Factors such as genetics, other health conditions and nutrition can contribute to gout.

Question 41

What drug is used to treat gout?

Answer 41

Allopurinol - it is a competitive inhibitor of XO so it decreases the amount of uric acid formed. Products of allopurinol treatment are Xanthine and hypoxanthine which are more soluble and less likely to form crystals.

Question 42

In addition to medication, how else can gout be controlled?

Answer 42

Diet - reduce ingestion of foods rich in purines

Question 43

Describe the pathway that recycles purines.

Answer 43

Adenine guanine and hypoxanthine are released from catabolism and recycled back to corresponding nucleotides

Adenine + PRPP —> AMP + PPi
(Catalyzed by Adenine phosphoribosyltransferase APRT)

Hypoxanthine + PRPP —> IMP+PPi
Guanine + PRPP —> GMP + PPi
(Both catalyzed by HGPRT)

Question 44

What is Lesch-Nyhan syndrome?

Answer 44

X-linked recessive genetic deficit in HGPRT, results in intellectual disability, aggression, and destructive behavior.

Lack of HGPRT leads to build up of PRPP which activates IMP synthesis, leads to increased [purines] and increased degradation and increased [uric acid]

Question 45

How are pyrimidines catabolized?

Answer 45

Nucleotides are catabolized to malonyl-CoA and release NH4+ in the process which is excreted as urea. The Malonyl-CoA can be used as a precursor to fatty acid synthesis or it can be made into succinyl CoA which is an intermediate of the CAC.