These are notes from lecture 9 of Harvard Extension’s biochemistry class.

amino acid catabolism

Amino acid catabolism generally accounts for about 10-15% of metabolic energy production. There are three circumstances where amino acids will undergo oxidative degradation / catabolism:

  1. Normal turnover of proteins
  2. When the diet is so protein-rich that it contains more amino acids than the body needs for protein synthesis
  3. During starvation or uncontrolled diabetes, glucose is unavailable and so amino acids are broken down instead

There are also three circumstances in which proteins are degraded into amino acids:

  1. To provide energy when needed (feeding into #3 from the list above)
  2. To eliminate abnormal proteins (see e.g. PrP lysosomal and proteasomal degradation)
  3. To regulate cellular processes (see e.g. cell cycle checkpoints)

There are only two major routes to protein degradation: lysosomes and proteasomes.  Lysosomal degradation includes endocytosis of extracellular waste, enclosure of intracellular waste, or chaperone-mediated autophagy.

Chaperone-mediated autophagy is important in the liver and kidney (the gluconeogenic organs) during starvation. Proteins with the KFERQ sequence (~30% of proteins in these organs) are selected. Usually these are rapid turnover cytosolic proteins including the glycolytic enzymes.   In this process, a chaperone such as Hsc70 recognizes the KFERQ targets those proteins to LAMP2A on the surface of lysosomes.  When the protein binds, LAMP2A multimerizes to act as a receptor which imports the protein into the lysosome for degradation.  The current thinking is that the KFERQ motif is usually hidden inside the protein and is exposed only when the protein partially unfolds under cellular stress.  But the main point of regulation in this process is LAMP2A expression level, which is upregulated during starvation.

Proteasomes degrade proteins tagged with at least 4 linked ubiquitins. Ubiquitin is a 76 residue protein present in all cells. Proteasomal degradation is involved in 1) normal turnover of metabolic proteins, 2) degrading defective proteins and 3) regulating processes such as the cell cycle. It’s not a major part of any starvation response.

Once proteins are degraded, the amino acids may be used in anabolism to make new proteins, or may be catabolized to produce energy.

Amino acid catabolism produces ammonia (NH3) and the carbon skeleton.  Ammonia is toxic and has to be managed by the urea cycle.  The carbon skeleton can have any one of four fates: CO2 + H2O, glucose, acetyl-CoA or ketone bodies.

Steps in amino acid catabolism

  1. transamination in which the amino group is transferred by an aminotransferase to α-ketoglutarate. After this is done you have α-keto acid and L-glutamate.
  2. The L-glutamate is de-aminated to yield α-ketoglutarate again. The ammonium goes to the urea cycle. α-ketoglutarate can be converted to glucose or can enter the citric acid cycle.

Steps 1 + 2 together are called transdeamination.

There are three ways to eliminate ammonia. Fish excrete ammonia directly. Birds and reptiles secrete uric acid. Terrestrial vertebrates mainly secrete urea (though we also secrete a small amount of uric acid in urine.)

The urea cycle takes place in the liver, the urea is secreted into the blood stream where the kidneys find it and excrete it in the urine.  Carbamoyl phosphate synthetase 1 creates carbamoyl phosphate which enters the cycle. Urea will ultimately be comprised of an NH3 from carbamoyl phosphate, an NH3 from aspartate and a C comes from bicarbonate (HCO3-).

When most tissues other than the liver produce ammonia, it is added to glutamate to form glutamine, and then glutamine can go into the bloodstream.  The exception is muscle, which has the glucose-alanine cycle, i.e. it uses alanine to hold the ammonia group to send it back to the liver.

The urea cycle is regulated a few ways. In the long term, prolonged starvation or high protein diets will result in upregulation of the enzymes involved in the cycle. In the short term, increased amino acid breakdown results in increased glutamate (due to transamination), which results in increased N-acetylglutamate, which allosterically activates carbamoyl phosphate synthetase 1.

Amino acids can be categorized as either glucogenic or ketogenic.  Glucogenic amino acids can be degraded to pyruvate, α-ketoglutarate, succinyl-CoA, fumarate or OAA all of which can feed into the citric acid cycle.  Ketogenic amino acids are degraded to acetyl-CoA and ketone bodies.

Phenylketonuria is a recessive disorder caused by loss of function of phenylalanine hydroxylase. Absent this enzyme, phenylalanine cannot be converted to tyrosine and so follows an alternate pathway leading to phenylpyruvate and then to phenylacetate. Phenylacetate imparts an odor to one’s urine. The disease phenotype is caused by elevated levels of both phenylalanine and its metabolites. Uncontrolled it causes mental retardation, but it can be well controlled by eating a bare minimum of phenylalanine-containing foods. You avoid aspartame (a DF dipeptide), eat very little protein, and supplement tyrosine (since it is normally produced from phenylalanine).

amino acid anabolism

Amino acids are synthesized from the carbon skeleton (derived from citric acid cycle intermediates) and from amino groups from glutamate or glutamine. We can synthesize 11 amino acids (nonessential). The other 9 are considered essential: H, I, L, K, M, F, T, W, V. Of these 9, some we cannot produce, others we can produce just not enough. Arginine is essential in children but adults can produce enough.

nucleotide anabolism

Nucleotides can be made two ways.  They can be made “from scratch” (“de novo synthesis”) from amino acids, CO2 and formate, or they can be “salvaged” from degradation products of other nucleic acids.

The de novo synthesis of purines involves many sources of atoms. Glycine, glutamine, aspartate, CO2 and formate all contribute.  Formate contributes in the form of formyltetrahydrofolate, where tetrahydrofolate (THF) is derived from dietary folic acid.  Ribose 5-phosphate for the backbone comes from the pentose phosphate pathway. It is converted to PRPP.  The first nucleotide reached is inosine monophosphate (IMP). From that you make AMP or GMP. The hydrolysis of ATP drives the synthesis of GMP and the hydrolysis of GTP drives the synthesis of AMP. After these are created, kinases let you reach ADP, GDP and GTP. Substrate level phosphorylation and oxidative phosphorylation let you reach ATP.

Purine synthesis is regulated several ways:

  1. ADP and GDP feed back and inhibit PRPP synthesis.
  2. All final nucleotides – ATP, ADP, AMP, GTP, GDP, GMP inhibit the first committed step of synthesis.
  3. AMP and GMP are competitive inhibitors against IMP, thus regulating their own production.
  4. Increased levels of PRPP activate production of AMP and GMP.

The de novo synthesis of pyrimidines follows an entirely separate path. Aspartate, glutamine, bicarbonate (HCO3-) and uridine monophosphate all contribute. In pyrimidine synthesis, UDP and UTP both feed back and inhibit carbamoyl phosphate synthetase 2.  This pathway yields UTP and CTP, but not TTP because so far we’ve talked only about ribonucleotide synthesis and not deoxyribonucleotide synthesis.

Once you have your ribonucleotides (ATP, GTP, CTP or UTP) these are converted to dNTP by ribonucleotide reductase.  Regulation of dNTP production is very important – obviously you can’t lack a nucleotide you need for DNA synthesis. On the flip side, if you have one in excess, it is also more likely to be inadvertently incorporated and cause a spontaneous DNA mutation. Thus excess and shortage are both big problems.

Hydroxyurea is a small molecule which scavenges a reactive tyrosyl radical (think tyrosine) at the active site of ribonucleotide reductase, inhibiting RNR and preventing the biosynthesis of dNTPs and therefore DNA replication. Therefore hydroxyurea is used as a chemotherapy drug. It’s also used as an anti-HIV drug, apparently with a few different mechanisms [reviewed in Lisziewicz 2003].

To obtain thymine, dUMP is converted to dTMP by thymidylate synthase using tetrahydrofolate from folic acid. Fluorouracil (FdUMP) is an irreversible inhibitor of thymidylate synthase which is used as a chemotherapy drug.  Folic acid is also a potential target to deplete dTTP and therefore inhibit rapidly dividing cells. There are also a whole class of anti-folate drugs (methotrexate, aminopterin, trimethoprim) which are competitive inhibitors of dihydrofolate reductase.

nucleic acid catabolism

Almost all dietary nucleic acid is degraded – therefore nucleic acid biosynthesis is very important. Nucleic acids from cellular turnover are also degraded.  Purines are catabolized from nucleotides into nucleosides, then into hypoxanthine and then xanthine oxidase (XO) converts hypoxanthine to xanthine and xanthine to uric acid. An accumulation of uric acid in the bloodstream and then joints causes gout. A treatment for gout is allopurinol, a xanthine oxidase inhibitor which causes hypoxanthine and xanthine to accumulate instead of uric acid.

Under conditions of cellular turnover, adenine guanine and hypoxanthine are released, and are recycled back to the corresponding nucleotides by phosphoribosyltransferases whihch add a PRPP group: adenine PPRT (APRT) and guanine-hypoxanthine PPRT (HGPRT). The X-linked genetic deficit of HGPRT causes PRPP to build up which causes mental retardation, aggressive and self-mutilating behavior in males.

Pyrimidine catabolism releases ammonia (converted to urea in the liver) and ends in malonyl-CoA, which is a precursor of fatty acid synthesis or can be converted into succinyl-CoA and go into the citric acid cycle.