1.

Nitrogen utilization
a. Amino acid catabolism
 Proteins are degraded into amino acids by action of proteolytic enzymes called Proteinases.
 The amino acids from various sources get mixed up to constitute a body called amino acid
pool.
 The amino acids in the pool have four fates:
o Be used for protein synthesis,
o Used in the synthesis of non-protein nitrogenous compounds e.g. hormones
o Decarboxylation
o Deamination.
 The first step in catabolism of most amino acids is transamination.
Involves the reversible transfer of amino group from amino acid to keto acid and is catalyzed by
aminotransferases.
 Keto acids are components of TCA cycle, e.g. oxaloacetate, α-ketoglutarate, pyruvate).
 Each amino acid except threonine and lysine has its own aminotransferase.
Common aminotransferases are aspartate aminotransferase, AST (also called serum
glutamateoxaloacetate-aminotransferase, SGOT) and alanine transaminase, ALT (also
called serum glutamate-pyruvate aminotransferase, SGPT).

 Almost all amino acids are transaminated to α-kg and glutamate to prepare them for
excretion.
 The amino group in the resultant alpha amino acid is converted to ammonia and excreted as
urea in urine.
 In all other tissues except the liver (e.g. skeletal muscle), glutamate is transaminated with
pyruvate to form alanine by alanine aminotransferase.
 This reaction provides an additional route for nitrogen transport from other tissues to the
liver.
 In the liver, alanine is transaminated with α-kg to regenerate pyruvate and glutamate.
 This process is referred to as the Glucose-alanine cycle.
 b. Non-essential amino acid biosynthesis
 In humans, only half of the standard amino acids (Glu, Gln, Pro, Asp, Asn, Ala, Gly, Ser,
Tyr, Cys) can be synthesized and are thus classified the nonessential amino acids.
 Within this group, glutamate, glutamine, and proline, have a shared anabolic pathway. It
begins with glutamate dehydrogenase, which adds ammonia to α-ketoglutarate in the
presence of NADPH to form glutamate. This is a key reaction for all amino acid synthesis:
glutamate is a nitrogen (amino group) donor for the production of all the other amino acids.
 Glutamine synthetase catalyzes the formation of glutamine from glutamate and ammonia.
This is an important biochemical reaction for a completely different reason: it is the primary
route for ammonia detoxification.
 Proline is synthesized from glutamate in a two-step process that begins with the reduction of
glutamate to a semialdehyde form that spontaneously cyclizes to D-pyrroline-5-carboxylate.
This is reduced by pyrroline carboxylate reductase to proline.
 Alanine and Aspartate are the products of glutamate-based transamination of pyruvate and
oxaloacetate, respectively.
 Asparagine is synthesized through one of two known pathways. In bacteria, an asparagine
synthetase combines aspartate and ammonia. However, in mammals, the aspartate gets its
amino group from glutamine.
 The synthesis of serine begins with the metabolic intermediate 3-phosphoglycerate
(glycolysis). Phosphoglycerate dehydrogenase oxidizes it to 3-phosphohydroxypyruvate. An
amino group is donated by glutamate in a reaction catalyzed by phosphoserine transaminase,
forming 3-phosphoserine, and finally the phosphate is removed by phosphoserine
phosphatase to produce serine.
 Serine is the immediate precursor to glycine, which is formed by serine
hydroxymethyltransferase. This enzyme requires the coenzyme tetrahydrofolate (THF),
which is a derivative of vitamin B9 (folic acid).
 Serine is also a precursor for cysteine, although the synthesis of cysteine actually begins with
the essential amino acid methionine. Methionine is converted to S-adenosylmethionine by
methionine adenosyltransferase. This is then converted to S-adenosylhomocysteine by a
member of the SAM-dependent methylase family. The sugar is removed by
adenosylhomocysteinase, and the resultant homocysteine is connected by cystathionine
synthase to the serine molecule to form cystathionine. Finally, cystathionine-g-lyase
catalyzes the production of cysteine.
 Tyrosine is another amino acid that depends on an essential amino acid as a precursor. In this
case, phenylalanine oxygenase reduces phenylalanine to produce the tyrosine.
Amino acid synthesis reactions can be grouped by biosynthetic families, named according to
their common starting metabolite.
α-Ketoglutarate Biosynthetic Family
Overview
Glutamate, glutamine, proline, and arginine are synthesized from α-ketoacids, which are
produced by the citric acid cycle.
Transamination is an essential reaction in the production of these amino acids.
The enzyme involved in this reaction is an aminotransferase.
Glutamate serves as a precursor for the production of many amino acids, such as glutamine,

α-ketoacid + glutamate ⇄ amino acid + α-ketoglutarate

proline, and arginine.

Glutamate itself is formed by amination of α-ketoglutarate: α-ketoglutarate + NH 4+ ⇄ glutamate

Glutamate synthesis
Glutamate is produced from α-ketoglutarate through the process of amination.
  α-ketoglutarate + NH4+ ⇄ glutamate
 Glutamate serves as the primary precursor for glutamine, proline, and arginine
Glutamine synthesis
Glutamate serves as the precursor for glutamine.
 Glutamate is converted to glutamine by glutamine synthetase.
 Glutamine synthetase uses:
o ATP
o NH4+: helps prevent ammonia build-up
 Glutamine synthetase inhibitors:
o Amino acids: glycine, alanine, serine, histidine, tryptophan
o Nucleotides: AMP, cytidine triphosphate (CTP)
o Other: carbamoyl phosphate, glucosamine-6-phosphate
 These inhibitors rely on glutamine for production, providing negative feedback when
glutamine levels are high.
 Glutamine synthetase activation:
 Glutamine synthetase is active in the deadenylated form.
 Adenylyl transferase is responsible for adenylation and de-adenylation.

Glutamine synthesis is mediated by glutamine synthetase: This reaction requires ATP.

Proline synthesis
Glutamate also serves as the precursor for proline.
3 steps to produce proline from glutamate:
1. Phosphorylation:
o Catalyzed by glutamate-5-kinase and glutamate-5-semialdehyde dehydrogenase
o Requires ATP and nicotinamide adenine dinucleotide (NADH)
o Produces glutamate-5-semialdehyde
2. Oxidation and dephosphorylation:
 Spontaneous cyclization reaction
 Produces 1-pyrroline-5-carboxylic acid
3. Reduction:
 Catalyzed by pyrroline-5-carboxylate reductase
 Requires NADH
 Generates proline
Cyclization and reduction reactions contributing to proline synthesis: These steps require ATP,
NADH, and a cyclization intermediate.
Arginine synthesis
Arginine is also derived from glutamate.
4 pathways to make arginine:
1. Reversible 2-step reaction
o From citrulline, aspartate, and ATP
2.  serves as an intermediate compound.
 Reversible demethylation from asymmetric dimethylarginine (ADMA)
 Reversible reaction from ornithine and urea
 Reversible reaction from citrulline, nitric oxide, and nicotinamide adenine dinucleotide
(NADP+)

3-Phosphoglycerate Biosynthetic Family

Serine, glycine, and cysteine are amino acids included in the 3-phosphoglycerate (3-PG)
biosynthetic family. Serine is the first amino acid produced in this group, and it subsequently
contributes to the production of glycine and cysteine.
Serine
Produced by a 3-step pathway:
1. Oxidation of 3-phosphoglycerate
o Catalyzed by phosphoglycerate dehydrogenase
o Produces 3-phosphohydroxypyruvate
o Inhibited by high concentrations of serine
 2. Transamination of 3-phosphohydroxypyruvate
o Catalyzed by phosphoserine transaminase
o Produces O-phosphoserine
3. Hydrolysis of O-phosphoserine
o Catalyzed by phosphoserine phosphatase
o Produces serine

Serine is made via 3 reactions, beginning with 3-phosphoglycerate (PG) and nicotinamide
adenine dinucleotide (NAD+)
Glycine
Glycine is produced from serine with the removal of a hydroxymethyl group.
 Catalyzed by serine hydroxymethyltransferase
 Requires tetrahydrofolate

Glycine is produced from serine: This 1-step reaction depends on the removal of a
hydroxymethyl group.
Cysteine
Cysteine is produced from serine and homocysteine through a 2-step pathway:
 1. Homocysteine and serine combine to form cystathionine
o Catalyzed by cystathionine beta-synthase
o Homocysteine contributes sulfur group
2. Cystathionine converted to cysteine
o Catalyzed by cystathionine gamma-lyase
o Also produces α-ketobutyrate

The formation of cysteine from serine and homocysteine

Oxaloacetate Biosynthetic Family
Overview
Aspartate, lysine, threonine, asparagine, methionine, and isoleucine are amino acids that have
oxaloacetate as a common precursor.
o Aspartate and asparagine are the only nonessential amino acids in this family.
o Oxaloacetate is first converted to aspartate.
o Aspartate is then converted to asparagine.
Aspartate
o Aspartate is produced by a transamination reaction.
o Catalyzed by an aminotransferase: transfers an amine group from another amino acid
o Produces aspartate from oxaloacetate.
Asparagine
Asparagine is also produced by a transamination reaction.
 Catalyzed by asparagine synthetase
 Produces asparagine from aspartate
Pyruvate Biosynthetic Family
Overview
Alanine, valine, and leucine share pyruvate as a common precursor. Alanine is the only amino
acid in the pyruvate biosynthetic family that can be produced by humans.
o Pyruvate is an end product of glycolysis
o Feedback inhibition is provided by the final product (e.g., alanine).
Alanine
Alanine is produced through a 2-step reaction.
1. α-ketoglutarate is converted to glutamate:
o Catalyzed by glutamate dehydrogenase
o Requires ammonia and NADH
2. Glutamate transfers an amino group to pyruvate to form alanine:
o Catalyzed by an aminotransferase enzyme
o Requires preformed pyruvate from glycolysis
Phosphoenolpyruvate Biosynthetic Family
Overview
The aromatic amino acids phenylalanine, tryptophan, and tyrosine are included in the
phosphoenolpyruvate family. Phenylalanine and tryptophan are essential amino acids; tyrosine is
a nonessential amino acid.
Tyrosine
Phenylalanine serves as the precursor for tyrosine
 The reaction is catalyzed by biopterin-dependent phenylalanine hydroxylase.
 Phenylalanine hydroxylase deficiency results in phenylketonuria.
 Tyrosine is the precursor for important neurotransmitters:
o Dopamine
o Noradrenaline
o Adrenaline
Clinical Relevance
 Phenylketonuria (PKU): defect of phenylalanine hydroxylase that results in impairment
of the conversion of phenylalanine to tyrosine and subsequent accumulation of
phenylalanine. Individuals will present with psychomotor delay and seizures, and their
 sweat will classically have a "mousy" odor. It is critical for these individuals to avoid
ingestion of phenylalanine.
 Maple syrup urine disease: defect in the branched-chain α-ketoacid dehydrogenase
complex that results in the accumulation of branched-chain amino acids. Individuals
present with cognitive disabilities, sweet-smelling urine, and dystonia. The primary
treatment is avoidance of branched-chain amino acids. Severe cases may require a liver
transplant.
 Homocystinuria: defect in the enzyme cystathionine β-synthase, which leads to an
accumulation of homocysteine. Individuals present with flushing, developmental delay,
downward lens dislocation, vascular disease, and osteoporosis. It is recommended that
these individuals maintain a diet low in sulfur.
 Alkaptonuria: deficiency of homogentisic acid dioxygenase, which impairs the normal
degradation of tyrosine to fumarate. Individuals present with a bluish-black discoloration
of connective tissues, arthritis, and calcifications of various tissues. There is currently no
treatment for alkaptonuria, but the life expectancy remains normal in these individuals.
c. Urea cycle
 Urea cycle is found primarily in liver and to a lesser extent in kidney.
 It converts the highly toxic ammonia to urea for excretion via urine.
 Amino acid catabolism produces urea which must be excreted.
 In the liver, glutamate undergoes oxidative deamination by action of L-glutamate
dehydrogenase in presence of NAD+ to form NH4+ and α-kg.
 The free NH4+ is immediately used together with CO2 generated in mitochondria to form
carbamoyl phosphate, a reaction catalyzed by carbamoyl synthetase 1 in presence of ATP.
 The carbamoyl phosphate enters the Urea (also called Krebs-Henseilet) cycle by
combining with ornithine.
 Urea is produced when arginine breaks down to regenerate ornithine.
Fig. 4.5: The urea cycle