Ammonium
0. 1 – 1.5 g
Buffering the urine
1
AMINO ACID METABOLISM
• Amino acids are required for the synthesis of proteins,
peptides, nucleotides, neurotransmitters, other amino acids
• Free amino acids can be provided to cells either from the
digestion of dietary proteins or the degradation of defective
or aged cellular proteins
• Amino acids are catabolized into components that can
directly join energy production pathways or be changed to
glucose, fatty acids or ketone bodies; this happens:
 when the amount of amino acids obtained from digestion
and degradation is more than what is needed for biosynthesis
 during starvation or uncontrolled diabetes mellitus
• The catabolism of amino acids produces:
 amino group – removed as urea
 carbon skeleton – seven types of intermediates
2
•
o
•
•
•
•
The digestion of proteins
The digestion of proteins begins in the stomach and
continues in the small intestine
Please refer to the notes on enzymes for the types, activation
and specificity of the proteases involved in the digestion
Rennin (chymosin) is important in infants because it breaks a
specific peptide bond in casein (a milk protein) curdling the
milk and increasing transit time in the stomach
Proteases can be endopeptidases or exopeptidases
The endopeptidases are specific for different types of
peptide bonds and produce fragments of varying sizes
Exopeptidases take over the job:
 carboxypeptidase A and B – produced by the pancreas and
cleave at the C-terminal
 brush border aminopeptidases – act on the N-terminal of
oligopeptides yielding free amino acids and di- and tripeptides
3
4
• Digestive enzymes themselves are digested and contribute
to the amino acid pool
• Free amino acids are transported into intestinal epithelial
cells by Na+ - dependent secondary transport
• Di- and tripeptides enter the epithelial cells through symport
with H+
 the H+ gradient is maintained by the Na+ - H+ exchanger
 peptidases in the epithelial cells change the di- and tripeptides into free amino acids
• Amino acids enter the portal vein by facilitated transport
• Many cells use Na+ - dependent secondary transport and to
some extent facilitated transport in order to absorb amino
acids
o premature activation of zymogens inside the pancreas
5
results in acute pancreatitis
6
•
•
•
•
•
•
Protein turnover
The degradation and resynthesis of proteins
The half-lives of eukaroytic proteins may vary from 30
seconds to many days
 ornithine decarboxylase – approx. 11 minutes;
hemoglobin – lifespan of red blood cells; crystallin (lens
protein) – life span of the organism
Rapidly degraded proteins include those proteins that are
defective due to wrong insertion of amino acids or damage
accumulated during normal functioning, regulatory enzymes,
A protein's half-life correlates with its N-terminal residue
Proteins with N-terminal Met, Ser, Ala, Thr, Val or Gly have
half lives greater than 20 hours
Proteins with N-terminal Phe, Leu, Asp, Lys or Arg have half
lives of 3 min or less
7
• PEST proteins having domains rich in Pro (P), Glu (E), Ser (S)
and Thr (T) are more rapidly degraded than other proteins
• There are two ways of intracellular degradation of proteins
1. lysosomal degradation - of endocytosed proteins or
proteins undergoing autophagy
• In autophagy, part of the cytoplasm may become
surrounded by two concentric membranes
• Fusion of the outer membrane of this autophagosome
with a lysosomal vesicle results in the degradation of
enclosed cytoplasmic structures and macromolecules
• the enzymes
responsible for the
degradation are
cathepsins
autophagosome
autophagic
vacuole
8
(lysosome)
2.
•
•
•
ATP- dependent cytosolic degradation
The ubiquitin – proteasome pathway
Ubiquitin is a small protein having 76 amino acids only
It is present in all eukaryotes (hence the name) and its
amino acid sequence is highly-conserved
• Ubiquitin marks proteins for death
 the carboxyl terminal of a ubiquitin forms an
isopeptide bond with the ε-amino group of a lysine
residue of a protein to be destroyed
• Three enzymes, are involved in the attachment of
ubiquitin:
Initially the terminal carboxyl group of ubiquitin is
joined in a thioester bond to a cysteine residue on
Ubiquitin-Activating Enzyme (E1) ; this is an ATPdependent process
9
 The ubiquitin is then transferred to a sulfhydryl
group on a Ubiquitin-Conjugating Enzyme (E2)
 Ubiquitin-Protein Ligase (E3) then promotes the
transfer of ubiquitin from E2 to the ε-amino group
of a Lys residue of a protein recognized by that E3
• The substrate-specificity of this system comes from
the various combinations of the types of E2 and E3
• More ubiquitins are added to form a polyubiquitin
chain
The terminal carboxyl of each ubiquitin is linked to
the ε-amino group of a lysine residue (Lys 29 or Lys
48) of the adjacent ubiquitin
 A chain of 4 or more ubiquitins targets proteins
for degradation in proteasomes
10
11
• The proteasome is a complex of multiple proteases
• Constitutes nearly 1 % of cellular protein
• It contains two main types of subcomplexes: a barrel-like
core particle (20S) and regulatory 19S particles on both
ends of the barrel
 the catalytic core particle and the regulatory
particles make up the functional 26S proteasome
• The 19 S particles may unfold proteins and translocate
the unfolded proteins into the 20 S particle; energy from
ATP is consumed in the process
• The 19 S particles also cleave isopeptide bonds and free
ubiquitin; ubiquitin is recycled
• The protein is degraded by the 20 S particle and free
amino acids are released into the cellular space
12
13
• Proteasomal degradation of particular proteins is an
essential mechanism by which cellular processes are
regulated, such as cell division, apoptosis,
differentiation and development
 progression through the cell cycle is controlled in
part through regulated degradation of proteins
called cyclins that activate cyclin-dependent kinases
• Inability to degrade proteins that activate cell division
(or rapid degradation of those proteins that suppress
tumor formation) can lead to cancer
• Diseases like Alzheimer's, Parkinson’s , type II
diabetes,… are associated with the deposition on
tissues of non-degradable protein aggregates known
as amyloid
14
The catabolism of amino acids
The fate of amino acid nitrogen
• α-amino groups are removed from amino acids mostly
through transamination reactions
 the amino acid becomes a keto acid when it donates
the amino group to α-ketoglutarate (changing it to
glutamate)
• All amino acids except lysine and threonine undergo
transamination reactions
• The enzymes involved are known as transaminases or
aminotransferases
• Pyridoxal phosphate (PLP) is the cofactor
• The glutamate thus derived collects the amino groups
and gives them off for biosynthesis or excretion
15
16
H
C
R
COO
EnzLysNH2
Enz
(CH2)4
NH2
Amino acid
HC
P
O
O
O

N
H
CH3
Enzyme (Lys)-PLP Schiff base
O
EnzLysNH2
CH2
O
H2
C
P
O
NH2
R
C

CH3
Pyridoxamine phosphate
H2
C
P
O
HC
O
O
OH

N
H
R
H
C
COO
N+
α-keto acid
O
CH3
Amino acid-PLP Shiff base (aldimine)
EnzLysNH2
COO
O
O
O

N
H
COO
H
H2
C
P

HC
O
O
H
H2
C
H
C
N+
N+
O
O
O
R
H
O
O

N
H
CH3
17
Amino acid-PLP Shiff base (aldimine)
• Free ammonium can be released in different ways:
A. The oxidative deamination of glutamate by glutamate
dehydrogenase - the only enzyme that can use NAD+ or
NADP+ as electron acceptor
The reaction is reversible and takes place in the
mitochondria
B. Serine and threonine dehydratases release NH4+;
histidine can be directly deaminated to give NH4+
C. Intestinal bacteria produce NH4+ from amino acids or
urea; the ammonia enters the portal vein
D. Glutamine and asparagine lose their side chain amino
groups through deamidation
E. The purine nucleotide cycle in the brain – aspartate is
used as a substrate and fumarate and ammonium are18
released
19
•
•
•
•
•
•
•
The urea cycle
Nitrogen balance is the difference between the amount of
nitrogen consumed and excreted per day
Positive nitrogen balance in growing individuals
Negative nitrogen balance during protein deficiency or
starvation
In healthy adults, the amount of nitrogen consumed and
excreted is approximately equal
Nitrogen is excreted in the form of urea, uric acid,
ammonium, creatinine, hippurate, creatine
Humans are ureotelic (excrete excess nitrogen mainly in the
form of urea)
 ammonotelic (ammonia); uricotelic (uric acid)
Ammonium made available in the liver from different
sources enters the urea cycle
20
• Since the enzymes of the urea cycle are present in the liver
only, amino groups from other tissues should be transported
to the liver
• Two mechanisms of transport:
1.the skeletal muscles export alanine synthesized from the
transamination of pyruvate (glucose catabolism) by
glutamate
 the amino group donated by glutamate was obtained from
the breakdown of amino acids in the muscle
21
2. Glutamate can accept another amino group through an ATPdependent reaction catalyzed by glutamine synthetase
 Glutamine is used by most tissues to transport ammonium
 Glutamine travels to the liver, kidneys and the intestine and
is deamidated
 the ammonium released by the deamidation is used as a
buffer (in the kidneys) or enters the urea cycle in the liver;
glutamine can be used as an energy source by the intestine
22
o The two nitrogen atoms of urea enter the urea cycle as NH4+
and as the amino N of aspartate
1. The synthesis of carbamoyl phosphate
• The NH4+ and HCO3- (carbonyl C) that will be part of urea are
incorporated first into carbamoyl phosphate
The cleavage of 2 ATP molecules is needed to form the
high energy carbamoyl phosphate
Carbamoyl phosphate synthetase (CPS I) is a
mitochondrial enzyme; the cytosolic isozyme is involved
in pyrimidine synthesis
• CPS I has an absolute requirement for the allosteric activator
N-acetylglutamate
• This derivative of glutamate is synthesized from acetyl-CoA
and glutamate when cellular glutamate is high, signaling an
excess of free amino acids due to protein breakdown or
23
dietary intake
2. The formation of citrulline
• Carbamoyl phosphate reacts with ornitihine to give citrulline;
catalyzed by ornithine transcarbamoylase
3. The entry of the second N
• Citrulline leaves the mitochondria in exchange for the entry of
ornithine from the cytosol
• Citrulline reacts with aspartate producing argininosuccinate
• Argininosuccinate synthetase requires the splitting of ATP to
24
AMP and PP
+
+
Pi
ornithine transcarbamoylase
ATP AMP+ PPi
argininosuccinate
synthetase
25
4. The formation of arginine
• Argininosuccinate lyase produces arginine and fumarate
• The arginine produced by the urea cycle is enough for adults
• The carbons of fumarate are those that were obtained from
aspartate
 Fumarate be changed to oxaloacetate by enzymes of the
citric acid cycle
 The oxaloacetate will receive an amino group from
glutamate and be changed to aspartate; aspartate
reenters the urea cycle
 The TCA and urea cycles constitute a bicycle: The Krebs
bicycle
5. The production of urea and the regeneration of ornithine
• The action of arginase produces urea and ornithine
26
• Urea travels to the kidneys and excreted through the urine
argininosuccinase
+
27
The urea cycle
28
The Krebs bicycle
The stocihiometry of the urea cycle
NH4++ CO2 + 3 ATP+ 2H2O→urea + fumarate + 2 ADP + AMP+ 4Pi
29
 In addition to the allosteric effects of N-acetylglutamate, the
synthesis of the enzymes of the urea cycle can be induced
during periods of increased metabolism – protein rich diet or
prolonged fasting
Urea cycle abnormalities
• Hereditary deficiency in any one of the urea cycle enzymes
or liver cirrhosis lead to the increase in the blood of
ammonia (hyperammonemia) or urea cycle intermediates
• The total lack of any urea cycle enzyme is lethal
• Elevated ammonia is toxic, especially to the brain
• Why is ammonia toxic to the brain? Hypotheses:
1. High ammonia levels would drive glutamine synthetase;
this would deplete glutamate – a neurotransmitter and
precursor for the synthesis of GABA
• Glutamine may exert osmotic effects leading to the
swelling of the brain
30
2. Depletion of glutamate and high ammonia levels would
drive the glutamate dehydrogenase reaction in the reverse
direction; the resulting depletion of α-ketoglutarate, inhibits
the production of energy
The treatment of urea cycle defects
• limiting protein intake to the amount barely adequate to
supply amino acids for growth, while adding to the diet the αketo acids of essential amino acids
• liver transplantation; gene therapy has also been tried
• If the defect occurs after the synthesis of argininosuccinate,
argininosuccinate can be used as a carrier for the removal of
nitrogen (because it has incorporated both amino groups)
 the problem in this situation would be one of
regenerating ornithine
 If arginase is not deficient, the intake of high amounts of
arginine would provide ornithine
31
• If the defect occurs at a point before the synthesis of
argininosuccinate, substances are used that form conjugates
with amino acids and are excreted in the urine
 The body has to use ammonia to replace the excreted
amino acids; ammonia levels decrease
 Drugs: Benzoic acid - reacts with glycine to give hippurate
Phenylbutyrate - first changed to phenylacetate
and then reacts with glutamine to produce
phenylacetylglutamine
• The most common defect is in ornithine transcarbamoylase
• In the rare cases of arginase deficiency, arginine should be
excluded from the diet
• Deficiency of N-acetyl glutamate can be corrected by
administering an analog, carbamoyl glutamate
32
33
•
•
•
•
•
•
The fate of the carbon skeleton of amino acids
The deamination of most amino acids yields α- keto acids
that, directly or via additional reactions, feed into major
metabolic pathways
Amino acids are grouped into two classes based on whether
or not their carbon skeletons can be converted to glucose:
glucogenic or ketogenic
Carbon skeletons of glucogenic amino acids are degraded to
pyruvate or a 4-C or 5-C intermediates of the Krebs cycle
Glucogenic amino acids are a major carbon source for
gluconeogenesis when glucose levels are low
They can also be catabolized for energy production or
converted to glycogen or fatty acids for energy storage
Carbon skeletons of ketogenic amino acids are degraded to
acetyl-CoA or acetoacetate
34
• Carbon skeletons of ketogenic amino acids can be
catabolized for energy or converted to ketone bodies or
fatty acids
• leucine and lysine are strictly ketogenic
• isoleucine, threonine, phenylalanine, tyrosine and
tryptophan and both glucogenic and ketogenic
• The remaining thirteen amino acids are glucogenic
amino acids producing pyruvate can be considered
ketogenic because pyruvate can be changed to acetyl-CoA
• One carbon transfer is a common theme in amino acid
metabolism
• Three cofactors are used to transfer different one carbon
groups between intermediates
Tetrahydrofolate (THF); S-adenosylmethionine/SAM/adoMet; Vitamin B 12 (5’-deoxyadensoyl/methyl cobalamin)
o Biotin is involved in the transfer of the most oxidized form of
35
carbon – CO2
36
•
•
•
•
•
•
Tetrahydrofolate
Folic acid/folate/folacin is composed of a pteridine nucleus, para
amino benzoic acid and one or more glutamic acid residues
Once folic acid is absorbed by the intestine, it is converted to
the biologically active form, tetrahydrofolate , by dihydrofolate
reductase
 only one glutamic acid remains; 4 hydrogens added
THF travels to the liver and glutamic acid residues are added
Most of the THF is released into the bile and recirculates just
like the bile acids
The carbon units carried by THF are attached to N5 and/or N10 of
the pteridine ring
One carbon units carried by THF are:
Most reduced: - CH3 (methyl)
Intermediate: - CH2 - (methylene)
Most oxidized: - CHO (formyl)
- CHNH (formimino)
37
- CH = (methenyl)
38
• The collection of one carbon units attached to THF is known
as the one – carbon pool
• While they are still attached to THF, the one carbon units can
be oxidized or reduced
• The main source of carbon units for THF is the carbon
removed during the conversion of serine to glycine
producing N5, N10-methylene THF
• Although THF can carry a methyl group at N5, the transfer
potential of the methyl group is insufficient for most
biosynthetic reactions; another cofactor is used as a carrier
of methyl groups
S-adenosyl methionine
• It contains an activated methyl thioether group; donates
methyl groups to oxygen or nitrogen
• Its synthesis to be discussed under the synthesis of cysteine
39
•
•
•
•
•
•
Vitamin B 12
The unmodified form of vitamin B 12 is known as cobalamin
It has a corrin ring similar to porphyrin
 but unlike porphyrin two of its pyrrole rings are joined
directly (no bridges); cobalt takes the place of iron
Cobalamin in the diet can be found in a free form or bound
with proteins
Free cobalamin is then bound by salivary or gastric secretions
known as haptocorrins; protein-bound cobalamin is first freed
of the proteins and then haptocorins bind it
Haptocorin bound cobalamin is changed to free cobalamin in
the intestine
Cobalamin is then bound by intrinsic factor which assists in
the absorption into the intestine; travels to the liver bound
with transcobalamin II
40
• Vitamin B 12 is involved in two reactions in the body:
1. The intramolecular rearrangement of a proton during the
formation of succinyl-CoA from propionyl-CoA
 the coenzyme form of vitamin B 12 used in this case is 5’- deoxy
-adenosyl cobalamin – 5’- deoxyadenosine attached to cobalt
2.The regeneration of methionine (to be discussed)
41
 methyl is attached to cobalt to give methyl cobalamin
X = 5’- deoxyadenosine or methyl
42
43
•
•
•
•
Essential and non-essential amino acids
Eleven amino acids are considered to be non-essential
because they can be synthesized in the body
arginine, cysteine, tyrosine and histidine are conditionally
essential
Arginine and histidine needed in the diet of children
and pregnant women; in adults arginine from the urea
cycle is enough and histidine is effectively recycled
 tyrosine and cysteine are synthesized from the
essential amino acids phenylalanine and methionine,
respectively; if these precursors are absent in the diet,
then the products become essential
The remaining essential amino acids are lysine, leucine,
isoleucine, valine, tryptophan, threonine and histidine
Except tyrosine and cysteine, essential amino acids can be
synthesized from glucose and ammonia (or another amino acid)
44
Amino acids related with intermediates of glycolysis
• In the synthesis of serine, 3-phosphoglycerate is sequentially
oxidized, transaminated and dephosphorylated
• In addition to the serine dehydratase reaction, serine can be
changed to pyruvate through transamination followed by
reduction and phosphorylation to give PEP (PEP then
changed to pyruvate)
• The main pathway of glycine synthesis is from serine – serine
hydroxymethyl transferase catalyzes the reaction which
involves PLP and N5, N10 methylene THF
 Glycine can be degraded by changing it to serine and then
to pyruvate
 A second way for the degradation of glycine is the
production of glyoxylate by D-amino acid oxidase
 D-amino oxidase is thought to act in the detoxification of
D-amino acids from bacteria or cooked foodstuffs
45
46
47
• Glyoxylate can react with α- ketglutarate and be directed
to energy production
• It can also be oxidized to oxalate by hepatic lactate
dehydrogenase
• This oxalate, along with the oxalate obtained from the
diet, contribute to the formation of kidney stones
 3/4th of kidney stones is composed of calcium oxalate
• The third and major approach to degrading glycine is by
glycine cleavage enzyme
Glycine degraded to NH4+, CO2 and –CH2– (carried by
THF)
If glycine cleavage enzyme is deficient, non-ketotic
hyperglycinemia results; mental retardation and early
death probably due to the increased inhibitory effects
of glycine on the nervous system
48
• The carbon skeleton and the amino group of cysteine are
derived from serine; the sulfur is transferred from methionine
• The sulfur of methionine attacks the 5’ carbon of the ribose
of ATP releasing all the three phosphates in the process;
SAM (Ado met) is the product
• SAM donates CH3 and becomes S-adenosylhomocysteine
• The hydrolytic removal of the adenosine gives homocysteine
• Serine reacts with homocysteine to give cystathionine –
catalyzed by cystathionine-β –synthase
• Cystathionine is cleaved by cystathionase to give cysteine
and α- ketobutyrate (which will be changed to propionylCoA and then succinyl-CoA)
• Homocysteine can be changed to methionine: N5-methyl
THF donates CH3 to cobalamin to give methyl cobalamin;
methyl cobalamin donates the CH3 to homocysteine and
methionine will be formed
49
50
• Cysteine allostericaly inhibits cystathionase and represses
the expression of the cystathionine -β-synthase gene
• The deficiency of cystathionine -β-synthase or poor binding
to its cofactor PLP would lead to increased amounts of
methionine and homocysteine in the blood
 Homocysteine would then dimerize to homocystine that
is excreted in the urine leading to homcystinuria
 excess homocysteine levels have been asssociated with
mental retardation and atherosclerosis (homocysteine
may damage the blood vessels and stimulate the
proliferation of smooth muscle cells)
• During the degradation of cysteine, the sulfur can be
disposed of in two ways
 the production of sulfuric acid
 the formation of PAPS (activated sulfur)
o The transamination of pyruvate yields alanine
51
Amino acids related to Krebs Cycle intermediates
1. α- ketoglutarate
• Glutamate by transamination or glutamate dehydrogenase
reaction
• From glutamate glutamine is synthesized by amidation;
glutamine synthetase is one of the only three enzymes of
humans that can fix free ammonia to organic molecules – the
other two enzymes are glutamate dehydrogenase and CPS I
• An intermediate known as glutamate γ semialdehyde is
synthesized from glutamate
• Proline and ornithine (which is precursor to arginine) can be
synthesized from or changed to glutamate and degraded
through glutamate γ semialdehyde
 if arginine is consumed in protein synthesis, more
ornithine would be synthesized from glutamate
52
53
54
• Although histidine is
effectively recycled in
humans, when degraded,
five of its carbons give rise
to glutamate
 Formiminoglutamate
obtained from histidine
is changed to glutamate
by transferring the
formimino group to THF
55
2. Oxaloacetate
• Oxaloacetate is transaminated to aspartate and aspartate
receives an amino group from glutamine and is changed to
asparagine
• Asparagine is broken down to aspartate and NH4+ by
asparaginase and aspartate is transaminated back to
oxaloacetate
3. Fumarate
• the urea cycle and the purine nucleotide cycle change the
56
carbons of aspartate to fumarate
•
Fumarate can be obtained from the breakdown of
tyrosine (meaning phenylalanine also can give fumarate)
4. Succinyl-CoA
• Methionine, threonine, valine and isoleucine are degraded
to propionyl-CoA that will be changed to succinyl-CoA
 methionine produces propionyl-CoA from αketobutyrate during the synthesis of cysteine
Threonine dehydratase also gives off α-ketobutyrate
• the main site of branched chain amino acid metabolism is
the muscles
• After transamination, the keto acids of all three amino
acids (valine, isoleucine and leucine undergo oxidative
decarboxylation; branched-chain α-keto acid
dehydrogenase complex – an analog of the PDC complex
57
• The subsequent reactions from all three amino acids give
the reduced equivalents NADH and FADH2
• Valine produces propionyl-CoA only
• Isoleucine degradation gives acetyl-CoA in addition to
propionyl-CoA
• leucine produces acetyl-CoA and acetoacetate (purely
ketogenic)
• Defects in branched-chain α-keto acid dehydrogenase
complex will lead to maple syrup urine disease; it can
progress to mental retardation
• Methylmalonyl CoA-mutase , an enzyme involved in the
processing of propionyl-CoA to succinyl-CoA, can be
deficient leading to methylmalonic acidemia (a rare but
deadly disorder)
58
59
Amino acids that form acetyl-CoA and acetoacetate
• Phenylalanine is hydroxylated to tyrosine by phenylalanine
hydroxylase, a mixed function oxidase
 Tetrahydrobiopterin (BH4) is the cofactor used
• Tyrosine is transaminated to p hydroxyphenylpyruvate
• p hydroxyphenylpyruvate is decarboxylated to homogentisate
by a dioxygenase
• Homogentisate 1,2- dioxygenase converts homogentisate to
maleylacetoacetate ; after two more steps, fumarate and
acetoacetate are obtained
• Phenylketonuria (PKU) results from the deficiency of
phenylalanine hydroxylase
 in this case, a minor pathway of phenylalanine metabolism,
transamination to give phenylpyruvate, becomes dominant
 phenylalanine and phenylpyuvate in the blood and urine
Mental retardation ensues; exclude phenylanine from diet 60
61
Tyrosinemia III
62
• homgentisate accumulates when homogentisate 1,2dioxygenase is deficient: alcaptonuria
Relatively benign; the urine turns black on standing
Later in life, the accumulation of homogentisate on the joints
may lead to arthritis
Sir Archibald Garrod pioneered the study of inborn errors of
metabolism based on his observations on alcaptonuria
• Tyrosinemias I-III may result from the deficiencies of the other
enzymes in tyrosine metabolism
• Tryptophan produces alanine from the non-ring carbons and
acetyl-CoA and formate from the ring structure
• The ring could also be used in the synthesis of NAD+ and
NADP+ - decreases the need for niacin in the diet
• A minor pathway of threonine degradation in the liver can
produce glycine and acetyl-CoA
• Lysine degradation produces acetyl-CoA and acetoacetate63
64
65
The relation between THF, Vit B12 and SAM
• N5-methyl THF is the most stable form of THF
• The only reaction in which methyl is removed from THF is
through transfer to vit B 12 during the synthesis of methionine
• If vit B 12 is deficient, N5-methyl THF would accumulate and
eventually most of the THF in the body would be found in the
form of N5-methyl THF - “Methyl Trap” hypothesis
 reactions that utilize folate would be compromised
• pernicious anemia has hematopoeitic and neurologic
components
• The hematopoeitic problems are thought to arise from a
secondary deficiency of folate resulting from the primary
deficiency in vit B 12 (absence of intrinsic factor)
• The neurologic disorders are caused by the absence of the
regenerating effect of vit B 12 on SAM; SAM is needed for
methylation reactions in the nervous tissue and also,
methylmalonyl-CoA competes with malonyl-CoA
66
67
Specialized products synthesized from amino acids
o Most neurostransmitters are either amino acids or derivatives of
amino acids
 Amino acids: glycine, glutamate, aspartate and γ aminobutyric acid (GABA)
• GABA is the most important inhibitory neurotransmitter in
the central nervous system
• It is synthesized through the decarboxylation of glutamate
• a characteristic feature of the production of biological amines
is decarboxylation which requires PLP
68
• Histidine is decarboxylated to histamine
Histamine is a mediator of allergic and other inflammatory
reactions, stimulator of gastric acid production and an
excitatory neurotransmitter in the brain
• Dopamine (D), epinephrine (E) and norepinephrine (NE) are
collectively known as catecholamines
 D and NE are excitatory neurotransmitters in the brain;
E and NE are also secreted by the adrenal medulla and
the peripheral nervous system
 the first step in the synthesis of catecholamines is the
BH4 dependent hydroxylation of tyrosine to form
3,4-dihydroxyphehnylalanine (DOPA)
 DOPA is then decarboxylated to dopamine
 Dopamine undergoes vitamin C dependent
hydroxylation to yield norepinephrine
 SAM methylates norepinephrine to epinephrine 69
70
• Parkinson’s disease is associated with low levels of dopamine
in the brain
 L- Dopa is used for treatment in the late stages of the
disease. Dopamine cannot cross the blood- brain barrier;
once L-Dopa gets into the brain, it will be changed to
dopamine
• The decarboxylation of tyrosine may produce tyramine which
binds to NE receptors and causes headaches and
hypertension if present in high quantities
 cheese, beer, red wine,… contain high amounts of
tyramine
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• Tryptophan is converted to the neurotransmitters
serotonin and melatonin in the pineal gland
• Tetrahydrobiopterin-dependent hydroxylation and PLPdependent decarboxylation are involved
• The conversion reactions are sensitive to light
• Serotonin accumulates in the brain during the daytime and
it is converted to N-acetylserotonin and then melatonin in
the dark
• Serotonin inhibits feeding and elevates the mood
Prozac, an anti-depressant acts by inhibiting serotonin
reuptake into the presynaptic neuron
• Melatonin may be involved in male sexual maturation; it
has roles in controlling the biological clock (cricadian
rhythm) and serves as an anti-0xidant
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• The catecholamines, tyramine and serotonin are
inactivated by oxidative deamination catalyzed by
monoamine oxidase (MAO) by methylation through the
action of catechol – O – methyl transferase (COMT)
• Histamine is first methylated by SAM and then acted upon
by MAO followed by another oxidation step
• GABA is inactivated by changing it back to glutamate and
then α-ketoglutatrate
• MAO produces H2O2 while degrading neurotransmitters
• MAO inhibitors are used in the early stages of Parkinson’s
disease and as antidepressants
 if people taking MAO inhibitors consume tryraminerich foods, the tyramine will not be degraded and this
will lead to serious hypertension – the “cheese effect”
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• Nitric oxide (NO) is produced from arginine
• Nitric oxide synthase (NOS) catalyzes a five electron oxdiation
of the guanidino nitrogen of arginine
• Two successive monoxygenation reactions occur to generate
the intermediate Nω–hydroxy-L-arginine
• NOS has got five prosthetic groups: FMN, FAD, heme, BH4 ,
and Ca2+ - calmodulin
• There are three tissue specific isozymes of NOS: neuronal
(nNOS), endothelial (eNOS) and inducible (iNOS)
• nNOS and eNOS activities are tightly regulated by Ca2+
• Ca2+ has no effect on iNOS, rather by cytokines; this is the
isoform that is involved in the production of the NO that is
produced by macrophages in order to kill microorganisms
• The NO produced by nNOS and eNOS acts in low
concentrations in the control of blood pressure,
neurotransmission, learning and memory
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• NO brings about vasodilation by activating guanylate cyclase
which increases the cellular level of cGMP
• Normally a phosphodiesterase terminates the action of NO
by changing cGMP to GMP
• Sildenafil (viagra) blocks a specific isozyme of phosphodiesterase and perpetuates the vasodilation
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o Amino acids are also used in the synthesis of substances
other than neurotransmitters
• Arginine and glycine are involved in the synthesis of creatine
• in the kidney, the guanidino group of arginine is transferred
to glycine to give ornithine and guanidoacetate
 in the liver, SAM methylates guanidoacetate to creatine
 ATP then donates a phosphate group to creatine to give
phosphocreatine
Creatine phosphate is used as a donor of phosphate in the
regeneration of ATP from ADP
Phoshpocreatine or creatine can be non-enzymatically
converted into the cyclic compound creatinine
• the urinary creatinine excretion of a person is extremely
constant from day to day and is proportional to the muscle
mass. 95 % of the creatine in the body is found in the skeletal
muscles and the remaining part in the heart, brain and testes
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• Creatinine Clearance Test: compares the level of creatinine in
urine (24 hrs.) with the creatinine level in the blood
• it is used for assessing kidney function
• Polyamines are positively charged molecules with multiple
amino groups that are found in high concentrations in cells
• The decarboxylation of ornithine produces putrescine
• Putrescine then reacts with decarboxylated SAM (from
methionine) and produces spermidine; spermidine is
changed by the same process into spermine
 Increase in the number of amino groups
• The decarboxylation of lysine and arginine would lead to
cadaverine and agmatine , respectively
• Polyamines may stabilize DNA by interacting with the
negatively charged phosphate groups of nucleotides
• They may also, to some extent, replace for cellular K+ and
Mg2+ and control nucleic acid and protein synthesis
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• Melanin is a family of polymeric pigments of different
colors synthesized from tyrosine
• tyrosinase catalyzes the conversion of tyrosine to DOPA
and DOPA to dopaquinone
• A number of intermediates follow from DOPA to finally
produce polymerized melanin
• Melanocytes produce melanin and carotene which blend
and give rise to the color of the skin, hair and eyes
• Melanin granules are uniformly distributed in melanocytes
and offer protection by absorbing ultra violet rays
• The deficiency of tyrosinase or other enzymes in the
synthesis pathway of melanin leads to albinism
 lack of pigment in the skin, eyes and hair; sensitivity to
sunlight
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