Nitrogen

Question 1

Amino acid pool

Answer 1

From

• dietary
  
  • stomach proteinases -> intestinal peptidases -> specific transporters
  • essential - must be in diet
• biosynthesis

Turnover with cellular proteins

Metabolism

• > biosynthesis (purines, pyrimidines, C1, neurotransmitters)
• > catabolism: C to gluconeogenesis, ATP; N to urea

Question 2

Regulation of protein synthesis

Answer 2

Synthesis -

• starvation -> (P) and sequestration of eIF2, eIF4 sequestered by 4EBP1
• insulin -> release eIF2, (P) of 4E-BP1 -> releases eIF4

Question 3

Amino acid catabolism

Answer 3

Aminotransferase
Amino acid + alpha-keto-glut alpha-keto-acid + glutamate

Specific for each amino acid - keto-acid pair
Ex:
 Alanine + aKG  pyruvate + Glut
 Aspartate + aKG  oxaloacetate + Glut
Usu with glutamate/alpha-ketoglutarate

Require pyridoxal (B6) (carries NH3)
Reversible, controlled by substrates (K = 1)

Question 4

Glutamate dehydrogenase

Answer 4

Oxidative deamination
Free amino acid -> amine transfer -> glutamate -> deamination to regenerate aKG

Glut + NAD NADH + NH3 + a-Ketoglutarate

Reversible (substrate dependent)
- ex post-prandial -> deamination
(-) ADP, GDP
(+) ATP, GTP

Question 5

Overview of urea cycle

Answer 5

Ammonia is toxic -> form urea
CO2 + NH4 + 3 ATP + Asp -> urea + 2ADP + AMP + fumarate
Irreversible due to pyrophosphate

Carbamoyl phos synthase I (CPS1) = rate limiting ->
carbamyl phos -(ornithine trans-carbamoylase) ->
citrulline -> transport out of mitochondria ->
citrulline + Asp -> argininosucc -(arg-succ lyase)>
Arg -(Arginase)> urea + ornithine -> transport back to mito
ornithin recycled via ornithin trans-carbamoylase

Question 6

Peripheral nitrogen

Answer 6

Alanine cycle (starvation)
Muscle:
 Pyruvate + NH3 -> Alanine ->
Liver:
 Alanine + aKG -> glutamate + pyruvate

Glutamine (2 NH3 groups)
Peripheral:
 Glutamate + NH4 +ATP -> glutamine
Liver:
 Glutamine -> glutamate + NH4

Question 7

Regulation of urea cycle

Answer 7

Carbamoyl phosphate synthetase I (CPS1)
(+) N-acetyl glutamate (NAG)
[(+) by overall amino acid levels, esp Arg]

Question 8

Protease regulation

Answer 8

Must be tightly controlled
Hydrolysis is energetically favorable - use ATP to prevent non-specific degradation

Proteases specific for substrate, cleavage location, different active site nucleophiles
Strategies:
 - natural protease inhibitors (serpins)
 - inactive precursors (zymogens)
 - compartmentalization (lysosomes)
 - specific signals (ubiquitin)

Question 9

Regulation of lysosomes

Answer 9

Fuse with endosome or autophagosome

Proteases = cathepsins

• compartmentalized
• need acidic conditions (proton pump)
• inhibited by cystatins in cytosol
• starvation -> inhibits mTOR kinase -> stimulates autophagy
• insulin -> Akt -> mTOR kinase -> inhibits

Question 10

Extracellular proteases

Answer 10

Ex:
tissue remodeling
complement, clotting, fibrinolysis
digestion

Serine proteases
Inhibited by “serpins” = “SERine Protease INbitor

Question 11

Ubiquitin pathway

Answer 11

Selective protein degradation

Ubiquitin (COOH-) -> thioester with E1 (requires ATP)

• > transfer to E2
• > combines with E3 = “ubiquitin ligase”
• > transfers to Lys amino groups on substrate
• > repeat until polyubiquitin chain

Proteasome = highly efficient
Receptor for ubiquitin -> de-ubiquitinating -> unfolding -> chaperone -> multiple proteases (different substrates)