Protein and Amino Acid Metabolism — Study Notes

1. Protein Digestion

Dietary proteins are too large to be absorbed directly and must be hydrolyzed into free amino acids, dipeptides, and tripeptides by proteases in the gastrointestinal tract.

Stomach

1. HCl: Secreted by parietal cells, it lowers gastric pH to ~1.5–2.5, denaturing dietary proteins (unfolding them to expose peptide bonds).
2. Pepsin: Secreted as the inactive zymogen pepsinogen by chief cells. Pepsinogen undergoes autocatalytic cleavage at low pH to form active pepsin. Pepsin is an endopeptidase that cleaves peptide bonds adjacent to aromatic residues (Phe, Tyr, Trp).

Small Intestine (Pancreatic Enzymes)

When acid chyme enters the duodenum, the pancreas secretes bicarbonate (neutralizing the acid) and inactive zymogens: - Enteropeptidase (an enzyme on the duodenal brush border) cleaves trypsinogen to form active trypsin. - Trypsin then cleaves and activates the other pancreatic zymogens: - Chymotrypsinogen $\rightarrow$ Chymotrypsin - Proelastase $\rightarrow$ Elastase - Procarboxypeptidases A and B $\rightarrow$ Carboxypeptidases A and B (exopeptidases cleaving from the C-terminus).

The resulting free amino acids, dipeptides, and tripeptides are absorbed by active transport systems (sodium-dependent cotransporters) into enterocytes, where any remaining small peptides are hydrolyzed to free amino acids before entering the portal circulation.

2. Amino Acid Catabolism (Deamination & Transamination)

Unlike lipids and carbohydrates, amino acids cannot be stored. Excess amino acids are degraded. The first step is the removal of the $\alpha$-amino group, leaving carbon skeletons (which are converted into glucose, lipids, or oxidized in the Citric Acid Cycle).

Transamination

This is the primary pathway for removing nitrogen. It involves the transfer of an amino group from an amino acid to an $\alpha$-keto acid (usually $\alpha$-ketoglutarate), converting the amino acid into its corresponding $\alpha$-keto acid and $\alpha$-ketoglutarate into glutamate:

$$\text{Amino Acid} + \alpha\text{-Ketoglutarate} \underset{\text{Aminotransferase}}{\overset{\text{PLP}}{\rightleftharpoons}} \alpha\text{-Keto Acid} + \text{Glutamate}$$

All aminotransferases (transaminases) require the coenzyme Pyridoxal Phosphate (PLP) (derived from Vitamin $B_6$) as a temporary amine carrier. Examples: - Alanine Aminotransferase (ALT): Alanine + $\alpha$-ketoglutarate $\rightleftharpoons$ Pyruvate + Glutamate. - Aspartate Aminotransferase (AST): Aspartate + $\alpha$-ketoglutarate $\rightleftharpoons$ Oxaloacetate + Glutamate.

Elevated serum ALT and AST are clinical markers of liver damage.

Oxidative Deamination

Glutamate collects nitrogen from transamination. In the liver, Glutamate Dehydrogenase (GDH) (located in the mitochondrial matrix) converts glutamate back to $\alpha$-ketoglutarate, releasing free ammonium ($NH_4^+$):

$$\text{Glutamate} + \text{NAD(P)}^+ + \text{H}_2\text{O} \xrightarrow{\text{GDH}} \alpha\text{-Ketoglutarate} + \text{NH}_4^+ + \text{NAD(P)H} + \text{H}^+$$

GDH is unique because it can utilize either $NAD^+$ (catabolic) or $NADP^+$ (anabolic).

graph LR
    A[Amino Acids] -->|Transamination / PLP| B[Glutamate]
    B -->|Glutamate Dehydrogenase| C[Ammonium NH4+]
    C -->|Urea Cycle| D[Urea - excreted in urine]

3. The Urea Cycle

Ammonium ($NH_4^+$) is highly toxic, particularly to the central nervous system, and must be eliminated. In humans, nitrogen is converted into urea in the liver and excreted by the kidneys. The Urea Cycle spans both the mitochondria and the cytosol.

The Five Steps

1. Synthesis of Carbamoyl Phosphate: $NH_4^+ + HCO_3^- + 2\text{ ATP} \rightarrow$ Carbamoyl Phosphate by Carbamoyl Phosphate Synthetase I (CPS-I) (mitochondrial). Rate-limiting step, obligatorily activated by N-acetylglutamate (NAG).
2. Synthesis of Citrulline: Carbamoyl phosphate + Ornithine $\rightarrow$ Citrulline by Ornithine Transcarbamoylase (mitochondrial). Citrulline is then transported to the cytosol.
3. Synthesis of Argininosuccinate: Citrulline + Aspartate (source of the second nitrogen atom of urea) + ATP $\rightarrow$ Argininosuccinate by Argininosuccinate Synthetase (cytosolic, consumes 2 ATP equivalents).
4. Cleavage to Arginine and Fumarate: Argininosuccinate $\rightarrow$ Arginine + Fumarate by Argininosuccinate Lyase (cytosolic). (Fumarate enters the Citric Acid Cycle, linking the two pathways).
5. Cleavage to Urea and Ornithine: Arginine + $H_2O \rightarrow$ Urea + Ornithine by Arginase (cytosolic). Ornithine is transported back into the mitochondria to start the cycle again.

Net Equation: $\text{NH}_4^+ + \text{HCO}_3^- + \text{Aspartate} + 3\text{ ATP} + \text{H}_2\text{O} \rightarrow \text{Urea} + \text{Fumarate} + 2\text{ ADP} + \text{AMP} + \text{PP}_i + 2\text{ P}_i$ (Cost = 4 high-energy bonds).

4. Essential and Non-Essential Amino Acids

Humans can synthesize only 10 of the 20 standard amino acids. The remaining 10 must be obtained from the diet.

• Essential Amino Acids (cannot be synthesized, must be ingested): Phenylalanine, Valine, Threonine, Tryptophan, Isoleucine, Methionine, Histidine, Arginine (essential in infants/during growth), Leucine, Lysine. (Mnemonic: "PVT TIM HALL").
• Non-Essential Amino Acids (can be synthesized by the body): Alanine, Asparagine, Aspartate, Glutamate, Glutamine, Glycine, Proline, Serine, Tyrosine (synthesized from Phe), Cysteine (gets sulfur from Met).

5. Biosynthesis of Non-Essential Amino Acids

Non-essential amino acids are synthesized from intermediates of glycolysis, the Citric Acid Cycle, or other amino acids: - From pyruvate: Alanine (via transamination). - From oxaloacetate: Aspartate (transamination) $\rightarrow$ Asparagine. - From $\alpha$-ketoglutarate: Glutamate (transamination or GDH) $\rightarrow$ Glutamine, Proline, Arginine. - From 3-phosphoglycerate (glycolysis): Serine $\rightarrow$ Glycine and Cysteine. - From Phenylalanine: Tyrosine (hydroxylase reaction, requires tetrahydrobiopterin). A defect in this enzyme causes Phenylketonuria (PKU).