Digestion of proteins. General pathways of amino acids
transformation.
Detoxification of
ammonia and biosynthesis of urea.
PROTEIN CATABOLISM –
hydrolysis breaks peptide bonds yielding amino acids
http://www.youtube.com/watch?v=SkkoE1RN_5E
AMINO ACID CATABOLISM -
(requires B6)
PROTEIN
CATABOLISM – attaches an amino group of an amino acid to a
keto acid converting a keto acid
into an amino acid. The original amino acid
becomes a keto acid.
1. New amino acid can be used for synthesis
2. Keto acid can be broken down in the TCA cycle
DEAMINATION – uses deaminase, water & NAD
1. breaks down an amino acid into a keto acid
and an ammonia.
2. liver cells convert ammonia to urea via the
PROTEIN ANABOLISM – dehydration synthesis
A. Amination – attaches amino group to a keto
acid
B. Ten essential amino acids
C. Deficiency diseases
1. marasmus
2. kwashiorkor
D. Genetic metabolic disorder - PKU
Breakdown of Food
and Fat
http://www.youtube.com/watch?v=AEsQxzeAry8
The
digestive process breaks down food by chemical and mechanical action into
substances that can pass into the bloodstream and be processed by body cells.
http://www.youtube.com/watch?v=g9G0zzdQx-M&feature=related
Certain
nutrients, such as salts and minerals, can be absorbed directly into the
circulation. Fat, complex carbohydrates, and proteins are broken down into
smaller molecules before being absorbed.
Fat
is split into glycerol and fatty acids; carbohydrates are split into
monosaccharide sugars; and proteins are split into linked amino acids called
peptides, and then into individual amino acids.
http://www.youtube.com/watch?v=STzOiRqzzL4
http://www.youtube.com/watch?v=tNdBdodTJNs&feature=related
http://www.youtube.com/watch?v=NewpaNwevFk
1: Mouth and oesophagus
Food
is chewed with the teeth and mixed with saliva. The enzyme amylase, present in saliva,
begins the breakdown of starch into sugar. Each lump of soft food, called a
bolus, is swallowed and propelled by contractions down the oesophagus into the
stomach.
2: Stomach
Pepsin
is an enzyme produced when pepsinogen, a substance secreted by the stomach
lining, is modified by hydrochloric acid (also produced by the stomach lining).
http://www.youtube.com/watch?v=1jtYH3RihcA
Pepsin
breaks proteins down into smaller units, called polypeptides and peptides.
Lipase is a stomach enzyme that breaks down fat into glycerol and fatty acids.
The acid produced by the stomach also kills bacteria.
3: Duodenum
Lipase,
a pancreatic enzyme, breaks down fat into glycerol and fatty acids. Amylase,
another enzyme produced by the pancreas, breaks down starch into maltose, a
disaccharide sugar. Trypsin and chymotrypsin are powerful pancreatic enzymes
that split proteins into polypeptides and peptides.
4: Small Intestine
Maltase,
sucrase, and lactase are enzymes produced by the lining of the small intestine.
They convert disaccharide sugars into monosaccharide sugars. Peptidase, another
enzyme produced in the intestine, splits large peptides into smaller peptides
and then into amino acids.
5: Large Intestine
Undigested
food enters the large intestine, where water and salt are absorbed by the
intestinal lining. The residue, together with waste pigments, dead cells , and
bacteria, is pressed into faeces and stored for excretion.
Urea Cycle
http://www.youtube.com/watch?v=AoBbVu5rnMs
Humans are
totally dependent on other organisms for converting atmospheric nitrogen into
forms available to the body. Nitrogen fixation is carried out by bacterial
nitrogensases forming reduced nitrogen, NH4+ which can
then be used by all organisms to form amino acids.
Overview of the flow of nitrogen in the biosphere. Nitrogen,
nitrites and nitrates are acted upon by bacteria (nitrogen fixation) and plants
and we assimilate these compounds as protein in our diets. Ammonia
incorporation in animals occurs through the actions of glutamate dehydrogenase
and glutamine synthase. Glutamate plays the central role in mammalian nitrogen
flow, serving as both a nitrogen donor and nitrogen acceptor.
Reduced nitrogen enters the human body as dietary free amino acids,
protein, and the ammonia produced by intestinal tract bacteria. A pair of
principal enzymes, glutamate dehydrogenase and glutamine synthatase, are found
in all organisms and effect the conversion of ammonia into the amino acids glutamate and glutamine,
respectively. Amino and amide groups from these 2 substances are freely
transferred to other carbon skeletons by transamination
and transamidation reactions.
Representative aminotransferase catalyzed reaction.
Aminotransferases exist for all amino acids except threonine and lysine.
The most common compounds involved as a donor/acceptor pair in transamination
reactions are glutamate and -ketoglutarate (-KG), which participate in reactions with many
different aminotransferases. Serum aminotransferases such as serum
glutamate-oxaloacetate-aminotransferase (SGOT) (also called aspartate
aminotransferase, AST) and serum glutamate-pyruvate
aminotransferase (SGPT) (also called alanine transaminase, ALT)
have been used as clinical markers of tissue damage, with increasing serum
levels indicating an increased extent of damage. Alanine transaminase
has an important function in the delivery of skeletal muscle carbon and
nitrogen (in the form of alanine) to the liver. In skeletal muscle, pyruvate is
transaminated to alanine, thus affording an additional route of nitrogen
transport from muscle to liver. In the liver alanine transaminase
tranfers the ammonia to -KG and regenerates pyruvate. The pyruvate can then be
diverted into gluconeogenesis. This process is refered to as the glucose-alanine cycle.
The Glutamate Dehydrogenase Reaction
The reaction catalyzed by glutamate dehydrogenase is:
NH4++-ketoglutarate+NAD(P)H+H+<---->glutamate+NAD(P)++H2O
Glutamate dehydrogenase can utilize either NAD orNADP
as cofactor. In the forward reaction as shown above glutamate
dehydrogenase is important in converting free ammonia and -ketoglutarate (-KG) to glutamate, forming one
of the 20 amino acids required for protein synthesis. However, it should be
recognized that the reverse reaction is a key anapleurotic process linking
amino acid metabolism with TCA cycle activity.In the reverse reaction, glutamate
dehydrogenase provides an oxidizable carbon source used for the
production of energy as well as a reduced electron carrier, NADH. As expected
for a branch point enzyme with an important link to energy metabolism, glutamate
dehydrogenase is regulated by the cell energy charge. ATP and GTP are
positive allosteric effectors of the formation of glutamate, whereas ADP and
GDP are positive allosteric effectors of the reverse reaction. Thus, when the
level of ATP is high, conversion of glutamate to -KG and other TCA cycle
intermediates is limited; when the cellular energy charge is low, glutamate is
converted to ammonia and oxidizable TCA cycle intermediates. Glutamate is also
a principal amino donor to other amino acids in subsequent transamination
reactions. The multiple roles of glutamate in nitrogen balance make it a
gateway between free ammonia and the amino groups of most amino acids.
The Glutamine Synthase Reaction
The reaction
catalyzed by glutamine synthase is:
glutamate +
NH4+ + ATP -------> glutamine + ADP + Pi +
H+
The glutamine
synthatase reaction is also important in several respects. First it
produces glutamine, one of the 20 major amino acids. Second, in animals,
glutamine is the major amino acid found in the circulatory system. Its role
there is to carry ammonia to and from various tissues but principally from
peripheral tissues to the kidney, where the amide nitrogen is hydrolyzed by the
enzyme glutaminase (reaction below); this process regenerates
glutamate and free ammonium ion, which is excreted in the urine.
glutamine + H2O -------> glutamate + NH3
Note that, in this function, ammonia arising in
peripheral tissue is carried in a nonionizable form which has none of the
neurotoxic or alkalosis-generating properties of free ammonia.
Liver
contains both glutamine synthetase and glutaminase
but the enzymes are localized in different cellular segments. This ensures that
the liver is neither a net producer nor consumer of glutamine. The differences
in cellular location of these two enzymes allows the liver to scavange ammonia
that has not been incorporated into urea. The enzymes of the urea cycle are
located in the same cells as those that contain glutaminase. The
result of the differential distribution of these two hepatic enzymes makes it
possible to control ammonia incorporation into either urea or glutamine, the
latter leads to excretion of ammonia by the kidney.
When acidosis occurs the body will divert more glutamine from the liver to the
kidney. This allows for the conservation of bicarbonate ion since the
incorporation of ammonia into urea requires bicarbonate (see below). When
glutamine enters the kidney, glutaminase releases one mole of
ammonia generating glutamate and then glutamate dehydrogenase
releases another mole of ammonia generating a-ketoglutarate. The ammonia will
ionizes to ammonium ion ( NH4+) which is excreted. The
net effect is a reduction in the pH (see also Kidneys and Acid-Base Balance).
While glutamine, glutamate, and the remaining nonessential amino acids can
be made by animals, the majority of the amino acids found in human tissues necessarily
come from dietary sources (about 400g of protein per day). Protein digestion
begins in the stomach, where a proenzyme called pepsinogen is
secreted, autocatalytically converted to Pepsin A, and used for
the first step of proteolysis. However, most proteolysis takes place in the
duodenum as a consequence of enzyme activities secreted by the pancreas. All of
the serine proteases and the zinc peptidases of pancreatic secretions are
produced in the form of their respective proenzymes. These proteases are both
endopeptidase and exopeptidase, and their combined action in the intestine
leads to the production of amino acids, dipeptides, and tripeptides, all of
which are taken up by enterocytes of the mucosal wall.
A circuitous regulatory pathway leading to the secretion of proenzymes into the
intestine is triggered by the appearance of food in the intestinal lumen.
Special mucosal endocrine cells secret the peptide hormones cholecystokinin (CCK) and secretin
into the circulatory system. Together, CCK and secretin cause contraction of
the gall bladder and the exocrine secretion of a bicarbonate-rich, alkaline
fluid, containing protease proenzymes from the pancreas into the intestine. A
second, paracrine role of CCK is to stimulate adjacent intestinal cells to
secrete enteropeptidase, a protease that cleaves trypsinogen
to produce trypsin. Trypsin also activates trypsinogen
as well as all the other proenzymes in the pancreatic secretion, producing the
active proteases and peptidases that hydrolyze dietary polypeptides.
Subsequent to luminal hydrolysis, small peptides and amino acids are
transferred through enterocytes to the portal circulation by diffusion,
facilitated diffusion, or active transport. A number of Na+-dependent
amino acid transport systems with overlapping amino acid specificity have been
described. In these transport systems, Na+ and amino acids at high
luminal concentrations are co-transported down their concentration gradient to
the interior of the cell. The ATP-dependent Na+/K+ pump
exchanges the accumulated Na+ for extracellular K+,
reducing intracellular Na+ levels and maintaining the high
extracellular Na+ concentration (high in the intestinal lumen, low
in enterocytes) required to drive this transport process.
Transport mechanisms of this nature are ubiquitous in the body. Small peptides
are accumulated by a proton (H+) driven transport process and
hydrolyzed by intracellular peptidases. Amino acids in the circulatory system
and in extracellular fluids are transported into cells of the body by at least
7 different ATP-requiring active transport systems with overlapping amino acid
specificities.
Hartnup disorder is an autosomal recessive impairment of neutral amino
acid transport affecting the kidney tubules and small intestine. It is believed
that the defect lies in a specific system responsible for neutral amino acid
transport across the brush-border membrane of renal and intestinal epithelium.
The exact defect has not yet been characterized. The characteristic diagnostic
feature of Hartnup disorder is a dramatic neutral hyperaminoaciduria.
Additionally, individuals excrete indolic compounds that originate from the
bacterial degradation of unabsorbed tryptophan. The reduced intestinal
absorption and increased renal loss of tryptophan lead to a reduced
availability of tryptophan for niacin and nicotinamide nucleotide biosynthesis.
As a consequence affected individuals frequently exhibit pellegra-like rashes.
.
Many other
nitrogenous compounds are found in the intestine. Most are bacterial products
of protein degradation. Some have powerful pharmacological (vasopressor)
effects.
|
Products of Intestinal Bacterial Activity |
||
|
Substrates |
Products |
|
|
- |
Vasopressor Amines |
Other |
|
Lysine |
Cadaverene |
- |
|
Arginine |
Agmatine |
- |
|
Tyrosine |
Tyramine |
- |
|
Ornithine |
Putrescine |
- |
|
Histidine |
Histamine |
- |
|
Tryptophan |
- |
Indole and
skatole |
|
All amino
acids |
- |
NH4+ |
Prokaryotes
such as E. coli can make the carbon skeletons of all 20 amino acids and
transaminate those carbon skeletons with nitrogen from glutamine or glutamate
to complete the amino acid structures. Humans cannot synthesize the branched
carbon chains found in branched chain amino acids or the ring systems found in
phenylalanine and the aromatic amino acids; nor can we incorporate sulfur into
covalently bonded structures. Therefore, the 10 so-called essential amino acids must be supplied from the diet.
Nevertheless, it should be recognized that,depending on the composition of the
diet and physiological state of an individual,one or another of the
non-essential amino acids may also become a required dietary component. For
example, arginine is not usually considered to be essential, because enough for
adult needs is made by the urea cycle.
However, the
urea cycle generally does not provide sufficient arginine for the needs of a
growing child.
To take a different type of example, cysteine and
tyrosine are considered non-essential but are formed from the essential amino
acids methionine and phenylalanine, respectively. If sufficient cysteine and
tyrosine are present in the diet, the requirements for methionine and
phenylalanine are markedly reduced; conversely, if methionine and phenylalanine
are present in only limited quantities, cysteine and tyrosine can become
essential dietary components. Finally, it should be recognized that if the -keto acids corresponding to the carbon skeleton of
the essential amino acids are supplied in the diet, aminotransferases in the
body will convert the keto acids to their respective amino acids, largely
supplying the basic needs.
Unlike fats and carbohydrates, nitrogen has no designated storage depots in the
body. Since the half-life of many proteins is short (on the order of hours),
insufficient dietary quantities of even one amino acid can quickly limit the
synthesis and lower the body levels of many essential proteins. The result of
limited synthesis and normal rates of protein degradation is that the balance
of nitrogen intake and nitrogen excretion is rapidly and significantly altered.
Normal, healthy adults are generally in nitrogen
balance, with intake and excretion being very well matched. Young
growing children, adults recovering from major illness, and pregnant women are
often in positive nitrogen balance. Their intake
of nitrogen exceeds their loss as net protein synthesis proceeds. When more
nitrogen is excreted than is incorporated into the body, an individual is in negative nitrogen balance. Insufficient quantities of
even one essential amino acid is adequate to turn an otherwise normal
individual into one with a negative nitrogen balance. The biological value of
dietary proteins is related to the extent to which they provide all the
necessary amino acids. Proteins of animal origin generally have a high
biological value; plant proteins have a wide range of values from almost none
to quite high. In general, plant proteins are deficient in lysine, methionine,
and tryptophan and are much less concentrated and less digestible than animal
proteins. The absence of lysine in low-grade cereal proteins, used as a dietary
mainstay in many underdeveloped countries, leads to an inability to synthesize
protein (because of missing essential amino acids) and ultimately to a syndrome
known as kwashiorkor, common among children in
these countries.
Essential vs. Nonessential Amino Acids
|
Nonessential |
Essential |
|
Alanine |
Arginine* |
|
Asparagine |
Histidine |
|
Aspartate |
Isoleucine |
|
Cysteine |
Leucine |
|
Glutamate |
Lysine |
|
Glutamine |
Methionine* |
|
Glycine |
Phenylalanine* |
|
Proline |
Threonine |
|
Serine |
Tryptophan |
|
Tyrosine |
Valine |
|
The amino acids arginine,
methionine and phenylalanine
are considered essential for reasons not directly related to lack of
synthesis. Arginine is synthesized by mammalian cells but at a rate that is
insufficient to meet the growth needs of the body and the majority that is
synthesized is cleaved to form urea. Methionine is required in large amounts
to produce cysteine if the latter amino acid is not adequately supplied in
the diet. Similarly, phenyalanine is needed in large amounts to form tyrosine
if the latter is not adequately supplied in the diet. |
|
Removal of Nitrogen
from Amino Acids
http://www.youtube.com/watch?v=5pBNunRmJn4
Nitrogen elimination begins intracellularly with protein
degradation. There are two main routes for converting intracellular proteins to
free amino acids: a lysosomal pathway, by which extracellular and some
intracellular proteins are degraded, and cytosolic pathways that are important
in degrading proteins of intracellular origin. In one cytosolic pathway a
protein known as ubiquitin is activated by conversion to an AMP derivative, and cytosolic
proteins that are damaged or otherwise destined for degradation are enzymically
tagged with the activated ubiquitin. Ubiquitin-tagged proteins are then
attacked by cytosolic ATP-dependent proteases that hydrolyze the targeted
protein, releasing the ubiquitin for further rounds of protein targeting.
The dominant reactions involved in removing amino acid nitrogen from the
body are known as transaminations. This class of
reactions funnels nitrogen from all free amino acids into a small number of
compounds; then, either they are oxidatively deaminated, producing ammonia, or
their amine groups are converted to urea by the urea cycle. Transaminations
involve moving an -amino group from a donor a-amino acid to the keto
carbon of an acceptor -keto acid. These reversible reactions are catalyzed
by a group of intracellular enzymes known as aminotransferases
(transaminases), which employ covalently bound pyridoxal phosphate as a
cofactor (see reaction mechanism).
Aminotransferases exist for all amino acids except
threonine and lysine. The most common compounds involved as a donor/acceptor
pair in transamination reactions are glutamic acid and -ketoglutaric acid, which participate in reactions
with many different aminotransferases. Serum aminotransferases such as serum
glutamate - oxaloacetate
- aminotransferase (SGOT) have been used as clinical markers of tissue
damage, with increasing serum levels indicating an increased extent of damage.
A small but clinically
important amount of creatinine is excreted in the urine daily, and the creatinine
clearance rate is often used as an indicator of kidney function. The
first reaction in creatinine formation is the transfer of the amido (or
amidine) group of arginine to glycine, forming guanidinoacetate. Subsequently,
a methyl group is transferred from the ubiquitous 1-carbon-donor S-adenosylmethionine
to guanidinoacetate to produce creatine (from which phosphocreatine is formed),
some of which spontaneously cyclizes to creatinine, and is eliminated in the
urine. The quantity of urine creatinine is generally constant for an individual
and approximately proportional to muscle mass. In individuals with damaged
muscle cells, creatine leaks out of the damaged tissue and is rapidly cyclized,
greatly increasing the quantity of circulating and urinary creatinine.
Because of the participation of -ketoglutarate in numerous transaminations,
glutamate is a prominent intermediate in nitrogen elimination as well as in
anabolic pathways. Glutamate formed in the course of nitrogen elimination is
either oxidatively deaminated by liver glutamate dehydrogenase,
forming ammonia, or converted to glutamine by glutamine synthase
and transported to kidney tubule cells. There the glutamine is sequentially
deamidated by glutaminase and deaminated by kidney glutamate
dehydrogenase.
The ammonia produced in the latter two reactions
is excreted as NH4+ in the urine, where it helps maintain
urine pH in the normal range of pH 4 to pH 8. The extensive production of
ammonia by peripheral or liver glutamate dehydrogenase is not
feasible because of the highly toxic effects of circulating ammonia. Normal
serum ammonium concentrations are in the range of 20-40 mmol, and an increase
in circulating ammonia to about 400 mmol causes alkalosis and neurotoxicity.
A final, therapeutically useful amino acid-related reaction is the amidation of
aspartic acid to produce asparagine. The enzyme asparagine synthase
catalyzes the ATP, requiring the transamidation reaction shown below:
aspartate+glutamine+ATP-------->glutamate+asparagine+AMP+PPi
Most cells perform this reaction well enough to produce all the asparagine
they need. However, some leukemia cells require exogenous asparagine, which
they obtain from the plasma. Chemotherapy using the enzyme asparaginase
takes advantage of this property of leukemic cells by hydrolyzing serum
asparagine to ammonia and aspartic acid, thus depriving the neoplastic cells of
the asparagine that is essential for their characteristic rapid growth.
In the peroxisomes of mammalian tissues, especially liver, there are 2 stereospecific
amino acid oxidases involved in elimination of amino acid
nitrogen. D-amino acid oxidase is an FAD-linked enzyme, and while
there are few D-amino acids that enter the human body the activity of this
enzyme in liver is quite high. L-amino acid oxidase is FMN-linked
and has broad specificity for the L amino acids.A number of substances,
including oxygen, can act as electron acceptors from the flavoproteins. If
oxygen is the acceptor the product is hydrogen peroxide, which is then rapidly
degraded by the catalases found in liver and other tissues. Missing or defective biogenesis of peroxisomes or L-amino acid
oxidase causes generalized hyper-aminoacidemia and hyper-aminoaciduria,
generally leading to neurotoxicity and early death.
The Urea
Cycle
http://www.youtube.com/watch?v=AoBbVu5rnMs&feature=related
http://www.youtube.com/watch?v=uFjfWexuMnM
Earlier it was noted that kidney glutaminase was responsible
for converting excess glutamine from the liver to urine ammonium. However,
about 80% of the excreted nitrogen is in the form of urea which is also largely
made in the liver, in a series of reactions that are distributed between the
mitochondrial matrix and the cytosol. The series of reactions that form urea is
known as the Urea Cycle or the Krebs-Henseleit Cycle.
http://www.youtube.com/watch?v=AoBbVu5rnMs&feature=related
Enzymes:
1: carbamoyl phosphate synthetase-I (CPS-I)
2: ornithine transcarbamoylase
(OTC)
3: argininosuccinate synthetase
4: argininosuccinase
5: arginase
http://www.youtube.com/watch?v=6VVyQkyK9_8&feature=related
The essential features of the urea cycle reactions and their metabolic
regulation are as follows: arginine from the diet or from protein breakdown is
cleaved by the cytosolic enzyme arginase, generating urea and
ornithine. In subsequent reactions of the urea cycle a new urea residue is
built on the ornithine, regenerating arginine and perpetuating the cycle.
Ornithine arising in the cytosol is transported to the mitochondrial matrix,
where ornithine transcabamoylase catalyzes the condensation of
ornithine with carbamoyl phosphate, producing citrulline. The energy for the
reaction is provided by the high-energy anhydride of carbamoyl phosphate.
The product, citrulline, is then transported to the cytosol, where the
remaining reactions of the cycle take place. The synthesis of citrulline
requires a prior synthesis of carbamoyl phosphate (CP).
The activation step requires 2 equivalents of ATP and the mitochondrial
matrix enzyme carbamoyl phosphate synthetase-I (CPS-I) (see reaction mechanism).There are two CP synthetases: a mitochondrial enzyme,
CPS-I, which forms CP destined for inclusion in the urea cycle, and a cytosolic
CP synthatase (CPS-II), which is involved in pyrimidine nucleotide biosynthesis. CPS-I is positively regulated by the allosteric
effector N-acetyl-glutamate, while the cytosolic
enzyme is acetylglutamate independent.
In a 2-step reaction, catalyzed by cytosolic argininosuccinate synthetase,
citrulline is converted to argininosuccinate.
The reaction involves the addition of AMP (from ATP) to the amido carbonyl
of citrulline, forming an activated intermediate on the enzyme surface
(AMP-citrulline), and the subsequent addition of aspartate to form
argininosuccinate.
Arginine and fumarate are produced from argininosuccinate by the cytosolic
enzyme argininosuccinate lyase.
In the final step of the cycle arginase cleaves urea from
aspartate, regenerating cytosolic ornithine, which can be transported to the
mitochondrial matrix for another round of urea synthesis.
Beginning and ending with ornithine, the reactions of the cycle consumes 3
equivalents of ATP and a total of 4 high-energy nucleotide phosphates. Urea is
the only new compound generated by the cycle; all other intermediates and
reactants are recycled.
The energy consumed in the production of urea is more than recovered by the
release of energy formed during the synthesis of the urea cycle intermediates.
Ammonia released during the glutamate dehydrogenase reaction is coupled
to the formation of NADH. In addition, when fumarate is converted back to
aspartate, the malate dehydrogenase reaction used to convert
malate to oxaloacetate generates a mole of NADH. These two moles of NADH, thus,
are oxidized in the mitochondria yielding 6 moles of ATP.
http://www.youtube.com/watch?v=uFjfWexuMnM
Regulation of
the Urea Cycle
The urea
cycle operates only to eliminate excess nitrogen. On high-protein diets the carbon
skeletons of the amino acids are oxidized for energy or stored as fat and
glycogen, but the amino nitrogen must be excreted. To facilitate this process,
enzymes of the urea cycle are controlled at the gene level. With long-term
changes in the quantity of dietary protein, changes of 20-fold or greater in
the concentration of cycle enzymes are observed. When dietary proteins increase
significantly, enzyme concentrations rise. On return to a balanced diet, enzyme
levels decline. Under conditions of starvation, enzyme levels rise as proteins
are degraded and amino acid carbon skeletons are used to provide energy, thus
increasing the quantity of nitrogen that must be excreted.
Short-term regulation of the cycle occurs principally at CPS-I,
which is relatively inactive in the absence of its allosteric activator N-acetylglutamate. The steady-state concentration of
N-acetylglutamate is set by the concentration of its components acetyl-CoA and
glutamate and by arginine, which is a positive allosteric effector of N-acetylglutamate
synthetase (glutamate transacylase).
A complete
lack of any one of the enzymes of the urea cycle will result in death shortly
after birth. However, deficiencies in each of the enzymes of the urea cycle,
including N-acetylglutamate synthase, have been identified. These
disorders are referred to as urea cycle disorders or
UCDs. More information on the individual UCDs
can be found in the Inborn Errors in Metabolism pages. A common thread to most UCDs is hyperammonemia leading to ammonia intoxication with
the consequences described below. Deficiencies in arginase
do not lead to symptomatic hyperammonemia as severe or as commonly as in the other
UCDs.
Clinical symptoms are most severe when the UCD is at the level of carbamoyl
phosphate synthetase I. Symptoms of UCDs usually arise at birth and
encompass, ataxia, convulsions, lethargy, poor feeding and eventually coma and
death if not recognized and treated properly. In fact, the mortality rate is
100% for UCDs that are left undiagnosed. Several UCDs manifest with late-onset
such as in adulthood. In these cases the symptoms are hyperactivity,
hepatomegaly and an avoidance of high protein foods.
In general, the treatment of UCDs has as common elements the reduction of
protein in the diet, removal of excess ammonia and replacement of intermediates
missing from the urea cycle. Administration of levulose
reduces ammonia through its action of acidifying the colon. Bacteria metabolize
levulose to acidic byproducts which then promotes excretion of ammonia in the
feces as ammonium ions, NH4+. Antibiotics can be administered to kill
intestinal ammonia producing bacteria. Sodium benzoate and sodium phenylbutyrate
can be administered to covalently bind glycine (forming hippurate) and
glutamine (forming phenylacetylglutamine), respectively. These latter
compounds, which contain the ammonia nitrogen, are excreted in the feces.
Dietary supplementation with arginine or citrulline can increase the rate of
urea production in certain UCDs.
|
UCD |
Enzyme
Deficiency |
Symptoms/Comments |
|
Carbamoylphosphate
synthetase I |
with 24h - 72h after birth infant becomes lethargic,
needs stimulation to feed, vomiting, increasing lethargy, hypothermia and
hyperventilation; without measurement of serum ammonia levels and appropriate
intervention infant will die: treament with arginine which activates
N-acetylglutamate synthetase |
|
|
N-acetylglutamate
synthetase |
severe hyperammonemia, mild hyperammonemia
associated with deep coma, acidosis, recurrent diarrhea, ataxia,
hypoglycemia, hyperornithinemia: treatment includes administration of
carbamoyl glutamate to activate CPS I |
|
|
Ornithine
transcarbamoylase |
most commonly occurring UCD, only X-linked UCD,
ammonia and amino acids elevated in serum, increased serum orotic acid due to
mitochondrial carbamoylphosphate entering cytosol and being incorporated into
pyrimidine nucleotides which leads to excess production and consequently
excess catabolic products: treat with high carbohydrate, low protein diet,
ammonia detoxification with sodium phenylacetate or sodium benzoate |
|
|
Argininosuccinate
synthetase |
episodic hyperammonemia, vomiting, lethargy, ataxia,
siezures, eventual coma: treat with arginine administration to enhance
citrulline excretion, also with sodium benzoate for ammonia detoxification |
|
|
Argininosuccinate
lyase |
episodic symptoms similar to classic citrullinemia,
elevated plasma and cerebral spinal fluid argininosuccinate: treat with
arginine and sodium benzoate |
|
|
Arginase |
rare UCD, progressive spastic quadriplegia and
mental retardation, ammonia and arginine high in cerebral spinal fluid and
serum, arginine, lysine and ornithine high in urine: treatment includes diet
of essential amino acids excluding arginine, low protein diet |
Neurotoxicity Associated with Ammonia
Earlier it
was noted that ammonia was neurotoxic. Marked brain damage is seen in cases of
failure to make urea via the urea cycle or to eliminate urea through the kidneys.
The result of either of these events is a buildup of circulating levels of
ammonium ion. Aside from its effect on blood pH, ammonia readily traverses the
brain blood barrier and in the brain is converted to glutamate via glutamate
dehydrogenase, depleting the brain of -ketoglutarate.
As the -ketoglutarate is depleted, oxaloacetate falls
correspondingly, and ultimately TCA cycle activity comes to a halt. In the
absence of aerobic oxidative phosphorylation and TCA cycle activity,
irreparable cell damage and neural cell death ensue. In addition, the increased
glutamate leads to glutamine formation. This depletes glutamate stores which
are needed in neural tissue since glutamate is both a neurotransmitter and a
precursor for the synthesis of -aminobutyrate, GABA, another neurotransmitter. Therefore, reductions in brain
glutamate affect energy production as well as neurotransmission.
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Introduction
All tissues have some capability for synthesis of the non-essential amino acids, amino acid remodeling, and conversion of
non-amino acid carbon skeletons into amino acids and other derivatives that
contain nitrogen. However, the liver is the major site of nitrogen metabolism in the body. In times of dietary surplus, the
potentially toxic nitrogen of amino acids is eliminated via transaminations,
deamination, and urea formation; the carbon skeletons are generally conserved
as carbohydrate, via gluconeogenesis, or as fatty acid via fatty acid synthesis pathways. In this respect amino acids fall into
three categories: glucogenic, ketogenic, or glucogenic and
ketogenic. Glucogenic amino acids are those that give rise to a net
production of pyruvate or TCA cycle intermediates, such as ketoglutarate or oxaloacetate, all of which are
precursors to glucose via gluconeogenesis. All amino acids except lysine and
leucine are at least partly glucogenic. Lysine and leucine are the only amino
acids that are solely ketogenic, giving rise only to acetylCoA or
acetoacetylCoA, neither of which can bring about net glucose production.
A small group of amino acids comprised of isoleucine,
phenylalanine, threonine, tryptophan, and tyrosine give rise to both glucose
and fatty acid precursors and are thus characterized as being glucogenic and
ketogenic. Finally, it should be recognized that amino acids have a third
possible fate. During times of starvation the reduced carbon skeleton is used
for energy production, with the result that it is oxidized to CO2
and H2O.
Amino Acid Biosynthesis
Glutamate
and Aspartate
Glutamate and aspartate are synthesized from their
widely distributed -keto acid precursors by simple 1-step transamination reactions. The former
catalyzed by glutamate dehydrogenase and the latter by aspartate
aminotransferase, AST.Aspartate is also derived from asparagine
through the action of asparaginase. The importance of glutamate as a
common intracellular amino donor for transamination reactions and of aspartate
as a precursor of ornithine for the urea cycle is described in the Nitrogen Metabolism page.
Alanine and the Glucose-Alanine Cycle
Aside
from its role in protein synthesis, alanine is second only to glutamine in
prominence as a circulating amino acid. In this capacity it serves a unique
role in the transfer of nitrogen from peripheral tissue to the liver. Alanine
is transferred to the circulation by many tissues, but mainly by muscle, in
which alanine is formed from pyruvate at a rate proportional to intracellular
pyruvate levels. Liver accumulates plasma alanine, reverses the transamination
that occurs in muscle, and proportionately increases urea production. The
pyruvate is either oxidized or converted to glucose via gluconeogenesis. When alanine transfer from muscle to liver is
coupled with glucose transport from liver back to muscle, the process is known
as the glucose-alanine cycle. The key feature of the cycle is that in 1
molecule, alanine, peripheral tissue exports pyruvate and ammonia (which are
potentially rate-limiting for metabolism) to the liver, where the carbon
skeleton is recycled and most nitrogen eliminated.There are 2 main pathways to
production of muscle alanine: directly from protein degradation, and via the
transamination of pyruvate by glutamate-pyruvate aminotransferase
(also called alanine transaminase, ALT).
glutamate + pyruvate <-------> -KG + alanine
The sulfur for cysteine synthesis comes from the
essential amino acid methionine. A condensation of ATP and methionine catalyzed
by methionine adenosyltransferase yields S-adenosylmethionine (SAM or AdoMet).
Biosynthesis of S-adenosylmethionine, SAM
SAM
serves as a precurosor for numerous methyl transfer reactions (e.g. the
conversion of norepinephrine to epinenephrine, see Specialized Products of Amino Acids). The result of methyl transfer is the
conversion of SAM to S-adenosylhomocysteine. S-adenosylhomocysteine is then
cleaved by adenosylhomocyteinase to yield homocysteine and adenosine.
Homocysteine can be converted back to methionine by methionine synthase,
a reaction that occurs under methionine-sparing conditions and requires N5-methyl-tetrahydrofolate
as methyl donor. This reaction was discussed in the context of vitamin B12-requiring
enzymes in the Vitamins page. Transmethylation reactions employing SAM are
extremely important, but in this case the role of S-adenosylmethionine in
transmethylation is secondary to the production of homocysteine (essentially a by-product
of transmethylase activity). In the production of SAM all phosphates of
an ATP are lost: one as Pi and two as PPi. It is
adenosine which is transferred to methionine and not AMP. In cysteine
synthesis, homocysteine condenses with serine to produce cystathionine, which
is subsequently cleaved by cystathionase to produce cysteine and -ketobutyrate. The sum of the latter two
reactions is known as trans-sulfuration. Cysteine is used for protein
synthesis and other body needs, while the ketobutyrate is decarboxylated and converted to
propionyl-CoA. While cysteine readily oxidizes in air to form the disulfide
cystine, cells contain little if any free cystine because the ubiquitous
reducing agent, glutathione effectively reverses the formation of
cystine by a non-enzymatic reduction reaction.
Utilization of methionine in the synthesis of
cysteine
The
2 key enzymes of this pathway, cystathionine synthase and cystathionase
(cystathionine lyase), both use pyridoxal phosphate as a cofactor, and both
are under regulatory control. Cystathionase is under negative allosteric
control by cysteine, as well, cysteine inhibits the expression of the cystathionine
synthase gene. Genetic defects are known for both the synthase and the
lyase. Missing or impaired cystathionine synthase leads to homocystinuria and is often associated with mental
retardation, although the complete syndrome is multifaceted and many
individuals with this disease are mentally normal. Some instances of genetic
homocystinuria respond favorably to pyridoxine therapy, suggesting that in
these cases the defect in cystathionine synthase is a decreased affinity
for the cofactor. Missing or impaired cystathionase leads to excretion
of cystathionine in the urine but does not have any other untoward effects.
Rare cases are known in which cystathionase is defective and operates at
a low level. This genetic disease leads to methioninuria with no other
consequences.
Tyrosine Biosynthesis
Tyrosine
is produced in cells by hydroxylating the essential amino acid phenylalanine.
This relationship is much like that between cysteine and methionine. Half of
the phenylalanine required goes into the production of tyrosine; if the diet is
rich in tyrosine itself, the requirements for phenylalanine are reduced by
about 50%. Phenylalanine hydroxylase is a mixed-function oxygenase:
one atom of oxygen is incorporated into water and the other into the hydroxyl
of tyrosine. The reductant is the tetrahydrofolate-related cofactor tetrahydrobiopterin, which is maintained in the reduced state by the
NADH-dependent enzyme dihydropteridine reductase.
Biosynthesis of tyrosine from phenylalanine
Missing
or deficient phenylalanine hydroxylase leads to the genetic
disease known as phenlyketonuria (PKU), which if untreated leads to severe mental
retardation. The mental retardation is caused by the accumulation of
phenylalanine, which becomes a major donor of amino groups in aminotransferase
activity and depletes neural tissue of -ketoglutarate. This absence of -ketoglutarate in the brain shuts down the TCA cycle and the associated production of aerobic energy,
which is essential to normal brain development.
The
product of phenylalanine transamination, phenylpyruvic acid, is reduced to
phenylacetate and phenyllactate, and all 3 compounds appear in the urine. The
presence of phenylacetate in the urine imparts a "mousy" odor. If the
problem is diagnosed early, the addition of tyrosine and restriction of phenylalanine
from the diet can minimize the extent of mental retardation.
In
other pathways, tetrahydrobiopterin is a cofactor. The effects of missing or
defective dihydropteridine reductase cause even more severe
neurological difficulties than those usually associated with PKU caused by
deficient hydroxylase activity.
Ornithine and Proline Biosynthesis
Glutamate
is the precursor of both proline and ornithine, with glutamate semialdehyde
being a branch point intermediate leading to one or the other of these 2
products. While ornithine is not one of the 20 amino acids used in protein
synthesis, it plays a significant role as the acceptor of carbamoyl phosphate
in the urea cycle. Ornithine serves an additional important role as the
precursor for the synthesis of the polyamines. The production of ornithine from glutamate is
important when dietary arginine, the other principal source of ornithine, is
limited. The fate of glutamate semialdehyde depends on prevailing cellular
conditions. Ornithine production occurs from the semialdehyde via a simple
glutamate-dependent transamination, producing ornithine.
Ornithine synthesis from glutamate
When
arginine concentrations become elevated, the ornithine contributed from the urea cycle plus that from glutamate semialdehyde inhibit
the aminotransferase reaction, with accumulation of the semialdehyde as a
result. The semialdehyde cyclizes spontaneously to 1pyrroline-5-carboxylate which is then reduced to proline by an
NADPH-dependent reductase.
The
main pathway to serine starts with the glycolytic intermediate
3-phosphoglycerate.
An
NADH-linked dehydrogenase converts 3-phosphoglycerate into a keto acid,
3-phosphopyruvate, suitable for subsequent transamination. Aminotransferase
activity with glutamate as a donor produces 3-phosphoserine, which is converted
to serine by phosphoserine phosphatase.
The
main pathway to glycine is a 1-step reaction catalyzed by serine
hydroxymethyltransferase.
This
reaction involves the transfer of the hydroxymethyl group from serine to the
cofactor tetrahydrofolate (THF), producing glycine and N5,N10-methylene-THF.
Glycine produced from serine or from the diet can also be oxidized by glycine
cleavage complex, GCC, to yield a second equivalent of N5,N10-methylene-tetrahydrofolate
as well as ammonia and CO2.
Glycine
is involved in many anabolic reactions other than protein synthesis including
the synthesis of purine nucleotides, heme, glutathione, creatine and serine.
Aspartate/Asparagine
and Glutamate/Glutamine Biosynthesis
Glutamate
is synthesized by the reductive amination of -ketoglutarate catalyzed by glutamate
dehydrogenase; it is thus a nitrogen-fixing reaction. In addition,
glutamate arises by aminotransferase reactions, with the amino nitrogen being
donated by a number of different amino acids. Thus, glutamate is a general
collector of amino nitrogen. Aspartate is formed in a transamintion reaction
catalyzed by aspartate transaminase, AST. This reaction uses the
aspartate -keto acid analog, oxaloacetate, and glutamate
as the amino donor. Aspartate can also be formed by deamination of asparagine
catalyzed by asparaginase. Asparagine synthetase
and glutamine synthetase, catalyze the production of asparagine
and glutamine from their respective amino acids. Glutamine is produced from
glutamate by the direct incorporation of ammonia; and this can be considered
another nitrogen fixing reaction. Asparagine, however, is formed by an
amidotransferase reaction. Aminotransferase reactions are readily reversible.
The direction of any individual transamination depends principally on the
concentration ratio of reactants and products. By contrast, transamidation
reactions, which are dependent on ATP, are considered irreversible. As a
consequence, the degradation of asparagine and glutamine take place by a
hydrolytic pathway rather than by a reversal of the pathway by which they were
formed. As indicated above, asparagine can be degraded to aspartate.
Amino Acid Catabolism
Glutamine/Glutamate and Asparagine/Aspartate
Catabolism
Glutaminase is an important kidney tubule enzyme involved in
converting glutamine (from liver and from other tissue) to glutamate and NH3+,
with the NH3+ being excreted in the urine. Glutaminase
activity is present in many other tissues as well, although its activity is not
nearly as prominent as in the kidney. The glutamate produced from glutamine is
converted to -ketoglutarate, making glutamine a glucogenic
amino acid. Asparaginase is also widely distributed within the
body, where it converts asparagine into ammonia and aspartate. Aspartate
transaminates to oxaloacetate, which follows the gluconeogenic pathway to
glucose. Glutamate and aspartate are important in collecting and eliminating
amino nitrogen via glutamine synthetase and the urea cycle, respectively. The catabolic path of the carbon
skeletons involves simple 1-step aminotransferase reactions that directly
produce net quantities of a TCA cycle intermediate. The glutamate dehydrogenase
reaction operating in the direction of -ketoglutarate production provides a second
avenue leading from glutamate to gluconeogenesis.
Alanine is also important in intertissue nitrogen
transport as part of the glucose-alanine cycle. Alanine's catabolic pathway involves a simple
aminotransferase reaction that directly produces pyruvate. Generally pyruvate
produced by this pathway will result in the formation of oxaloacetate, although
when the energy charge of a cell is low the pyruvate will be oxidized to CO2
and H2O via the PDH complex and the TCA cycle. This makes alanine a glucogenic amino acid.
Arginine,
Ornithine and Proline Catabolism
The catabolism of arginine begins within the context
of the urea cycle. It is hydrolyzed to urea and ornithine by arginase.
Ornithine, in excess of urea cycle needs, is transaminated to form glutamate
semialdehyde. Glutamate semialdehyde can serve as the precursor for proline
biosynthesis as described above or it can be converted to glutamate. Proline
catabolism is a reversal of its synthesis process. The glutamate semialdehyde
generated from ornithine and proline catabolism is oxidized to glutamate by an
ATP-independent glutamate semialdehyde dehydrogenase. The
glutamate can then be converted to ketoglutarate in a transamination reaction. Thus
arginine, ornithine and proline, are glucogenic.
PAPS is used for the transfer of sulfate to biological
molecules such as the sugars of the glycosphingolipids.Other than protein, the most important product
of cysteine metabolism is the bile salt precursor taurine, which is used to form the bile acid conjugates taurocholate and taurochenodeoxycholate.The enzyme cystathionase can also
transfer the sulfur from one cysteine to another generating thiocysteine and
pyruvate. Transamination of cysteine yields -mercaptopyruvate which then reacts with
sulfite, (SO32-), to produce thiosulfate, (S2O32-)
and pyruvate. Both thiocysteine and thiosulfate can be used by the enzyme rhodanese
to incorporate sulfur into cyanide, (CN-), thereby detoxifying the
cyanide to thiocyanate.
The principal fates of the essential amino acid
methionine are incorporation into polypeptide chains, and use in the production
of -ketobutyrate and cysteine via SAM as described
above. The transulfuration reactions that produce cysteine from homocysteine
and serine also produce -ketobutyrate, the latter being converted to succinyl-CoA. Regulation of
the methionine metabolic pathway is based on the availability of methionine and
cysteine. If both amino acids are present in adequate quantities, SAM
accumulates and is a positive effector on cystathionine synthase,
encouraging the production of cysteine and ketobutyrate (both of which are glucogenic).
However, if methionine is scarce, SAM will form only in small quantities, thus
limiting cystathionine synthase activity. Under these conditions
accumulated homocysteine is remethylated to methionine, using N5-methylTHF
and other compounds as methyl donors.
Valine, Leucine
and Isoleucine Catabolism
This group of essential amino acids are identified as
the branched-chain amino
acids, BCAAs. Because
this arrangement of carbon atoms cannot be made by humans, these amino acids
are an essential element in the diet. The catabolism of all three compounds
initiates in muscle and yields NADH and FADH2 which can be utilized
for ATP generation. The catabolism of all three of these amino acids uses the
same enzymes in the first two steps. The first step in each case is a
transamination using a single BCAA aminotransferase, with -ketoglutarate as amine acceptor. As a result,
three different -keto acids are produced and are oxidized using a common branched-chain
-keto acid dehydrogenase, yielding the three different CoA derivatives.
Subsequently the metabolic pathways diverge, producing many intermediates. The principal
product from valine is propionylCoA, the glucogenic precursor of succinyl-CoA.
Isoleucine catabolism terminates with production of acetylCoA and propionylCoA;
thus isoleucine is both glucogenic and ketogenic. Leucine gives rise to
acetylCoA and acetoacetylCoA, and is thus classified as strictly ketogenic.
There are a number of genetic diseases associated with faulty catabolism of the
BCAAs. The most common defect is in the branched-chain -keto acid dehydrogenase. Since there is only one dehydrogenase enzyme
for all three amino acids, all three keto acids accumulate and are excreted in the
urine. The disease is known as Maple syrup urine disease because of the characteristic odor of the urine
in afflicted individuals. Mental retardation in these cases is extensive.
Unfortunately, since these are essential amino acids, they cannot be heavily
restricted in the diet; ultimately, the life of afflicted individuals is short
and development is abnormal The main neurological problems are due to poor
formation of myelin in the CNS.
Phenylalanine
and Tyrosine Catabolism
Phenylalanine normally has only two fates:
incorporation into polypeptide chains, and production of tyrosine via the
tetrahydrobiopterin-requiring phenylalanine hydroxylase. Thus,
phenylalanine catabolism always follows the pathway of tyrosine catabolism. The
main pathway for tyrosine degradation involves conversion to fumarate and
acetoacetate, allowing phenylalanine and tyrosine to be classified as both
glucogenic and ketogenic. Tyrosine is equally important for protein
biosynthesis as well as an intermediate in the biosynthesis of several
physiologically important metabolites e.g. dopamine, norepinephrine and epinephrine
(see Specialized Products of Amino Acids).
http://www.youtube.com/watch?v=hpaki7F4HR0
http://www.youtube.com/watch?v=CWfrVS4Bm1Y&feature=related
As in phenylketonuria (deficiency of phenylalanine
hydroxylase), deficiency of tyrosine transaminase leads
to urinary excretion of tyrosine and the intermediates between phenylalanine
and tyrosine. The adverse neurological symptoms are the same for the two
diseases. Genetic diseases (such as various tyrosinemias and alkaptonuria) are
also associated with other defective enzymes of the tyrosine catabolic pathway.
The first genetic disease ever recognized, alcaptonuria, is caused by defective homogentisic acid
oxidase. Homogentisic acid accumulation is relatively innocuous,
causing urine to darken on exposure to air, but no life-threatening effects
accompany the disease. The other genetic deficiencies lead to more severe
symptoms, most of which are associated with abnormal neural development, mental
retardation, and shortened life span.
Lysine catabolism is unusual in the way that the -amino group is transferred to -ketoglutarate and into the general nitrogen
pool. The reaction is a transamination in which the -amino group is transferred to the -keto carbon of -ketoglutarate forming the metabolite, saccharopine. Unlike the majority of transamination reactions,
this one does not employ pyridoxal phosphate as a cofactor. Saccharopine is
immediately hydrolyzed by the enzyme -aminoadipic semialdehyde synthase in such a way that the amino nitrogen remains
with the -carbon of -ketoglutarate, producing glutamate and -aminoadipic semialdehyde. Because this
transamination reaction is not reversible, lysine is an essential amino acid.
The ultimate end-product of lysine catabolism is acetoacetyl-CoA Genetic
deficiencies in the enzyme -aminoadipic semialdehyde synthase have been observed in individuals who excrete
large quantities of urinary lysine and some saccharopine. The lysinemia and
associated lysinuria are benign. Other serious disorders associated with lysine
metabolism are due to failure of the transport system for lysine and the other
dibasic amino acids across the intestinal wall. Lysine is essential for protein
synthesis; a deficiencies of its transport into the body can cause seriously
diminished levels of protein synthesis. Probably more significant however, is
the fact that arginine is transported on the same dibasic amino acid carrier,
and resulting arginine deficiencies limit the quantity of ornithine available
for the urea cycle. The result is severe hyperammonemia after a meal
rich in protein. The addition of citrulline to the diet prevents the
hyperammonemia. Lysine is also important as a precursor for the synthesis of carnitine, required for the transport of fatty acids into the
mitochondria for oxidation. Free lysine does not serve as the precursor for
this reaction, rather the modified lysine found in certain proteins. Some
proteins modify lysine to trimethyllysine using SAM as the methyl donor to transfer
methyl groups to the -amino of the lysine side chain. Hydrolysis of proteins containing
trimethyllysine provide the substrate for the subsequent conversion to
carnitine.
Histidine catabolism begins with release of the amino group catalyzed by histidase,
introducing a double bond into the molecule. As a result, the deaminated product,
urocanate, is not the usual -keto acid associated with loss of -amino nitrogens. The end product of histidine
catabolism is glutamate, making histidine one of the glucogenic amino acids.
Another key feature of histidine catabolism is that it serves as a source of
ring nitrogen to combine with tetrahydrofolate (THF), producing the 1-carbon
THF intermediate known as N5-formiminoTHF. The latter
reaction is one of two routes to N5-formiminoTHF. The
principal genetic deficiency associated with histidine metabolism is absence or
deficiency of the first enzyme of the pathway, histidase. The
resultant histidinemia is relatively benign. The disease, which is of
relatively high incidence (
Tryptophan Catabolism
A number of important side reactions occur during the
catabolism of tryptophan on the pathway to acetoacetate. The first enzyme of
the catabolic pathway is an iron porphyrin oxygenase that opens the indole
ring. The latter enzyme is highly inducible, its concentration rising almost
10-fold on a diet high in tryptophan. Kynurenine is the first key branch point intermediate in the
pathway. Kynurenine undergoes deamniation in a standard transamination reaction
yielding kynurenic acid. Kynurenic acid and metabolites have been shown
to act as antiexcitotoxics and anticonvulsives. A second side branch reaction
produces anthranilic acid plus alanine. Another equivalent of alanine is
produced further along the main catabolic pathway, and it is the production of
these alanine residues that allows tryptophan to be classified among the
glucogenic and ketogenic amino acids. The second important branch point
converts kynurenine into 2-amino-3-carboxymuconic semialdehyde, which has two
fates. The main flow of carbon elements from this intermediate is to glutarate.
An important side reaction in liver is a transamination and several
rearrangements to produce limited amounts of nicotinic acid, which leads to production of a small amount of NAD+
and NADP+ Aside form its role as an amino acid in protein
biosynthesis, tryptophan also serves as a precursor for the synthesis of serotonin and melatonin. These products are discussed in Specialized Products of Amino Acids
Specialized Products of Amino Acids
Tyrosine-Derived Neurotransmitters
The majority
of tyrosine that does not get incorporated into proteins is catabolized for
energy production. One other significant fate of tyrosine is conversion to the catecholamines. The catecholamine neurotransmitters
are dopamine, norepinephrine,
and epinephrine (see also Biochemistry of Nerve Transmission).Norepinephrine is the principal neurotransmitter of
sympathetic postganglionic endings. Both norepinephrine and the methylated
derivative, epinephrine are stored in synaptic knobs of neurons that secrete
it, however, epinephrine is not a mediator at postganglionic sympathetic
endings.Tyrosine is transported into catecholamine-secreting neurons and
adrenal medullary cells where catechaolamine synthesis takes place. The first
step in the process requires tyrosine hydroxylase, which like phenylalanine
hydroxylase requires tetrahydrobiopterin as cofactor. The hydroxylation
reaction generates DOPA (3,4-dihydrophenylalanine). DOPA decarboxylase
converts DOPA to dopamine, dopamine b-hydroxylase converts
dopamine to norepinephrine and phenylethanolamine N-methyltransferase
converts norepinephrine to epinephrine. This latter reaction is one of several
in the body that uses SAM as a methyl donor generating S-adenosylhomocysteine.
Within the substantia nigra and some other regions of the brain, synthesis
proceeds only to dopamine. Within the adrenal medulla dopamine is converted to
norepinephrine and epinephrine.
Synthesis of the catecholamines from tyrosine.
Once
synthesized, dopamine, norepinephrine and epinephrine are packaged in
granulated vesicles. Within these vesicles, norepinephrine and epinephrine are
bound to ATP and a protein called chromogranin A.
Metabolism of the catecholemines occurs through the actions of catecholamine-O-methyltransferase,
(COMT) and monoamine oxidase, (MAO). Both of these enzymes
are widley distributed throughout the body. However, COMT is not found in nerve
endings as is MAO.
Tryptophan-Derived
Neurotransmitters
Tryptopan
serves as the precursor for the synthesis of serotonin
(5-hydroxytryptamine, 5-HT, see also Biochemistry of Nerve Transmission) and melatonin
(N-acetyl-5-methoxytryptamine).
Serotonin is
synthesized through 2-step process involving a tetrahydrobiopterin-dependent
hydroxylation reaction (catalyzed by tryptophan-5-monooxygenase)
and then a decarboxylation catalyzed by aromatic L-amino acid
decarboxylase. The hydroxylase is normally not saturated and as a
result, an increased uptake of tryptophan in the diet will lead to increased
brain serotonin content. Serotonin is present at highest concentrations in
platelets and in the gastrointestinal tract. Lesser amounts are found in the
brain and the retina. Serotonin containing neurons have their cell bodies in
the midline raphe nuclei of the brain stem and project to portions of the
hypothalamus, the limbic system, the neocortex and the spinal cord. After
release from serotonergic neurons, most of the released serotonin is recaptured
by an active reuptake mechanism. The function of the antidepressant, Prozac is to inhibit this reuptake process,
thereby, resulting in prolonged serotonin presence in the synaptic cleft. The
function of serotonin is exerted upon its interaction with specific receptors.
Several serotonin receptors have been cloned and are identified as 5HT1,
5HT2, 5HT3, 5HT4, 5HT5, 5HT6,
and 5HT7. Within the 5HT1 group there are subtypes 5HT1A,
5HT1B, 5HT1D, 5HT1E, and 5HT1F.
There are three 5HT2 subtypes, 5HT2A, 5HT2B,
and 5HT2C as well as two 5HT5 subtypes, 5HT5a
and 5HT5B. Most of these receptors are coupled to G-proteins that
affect the activities of either adenylate cyclase or phospholipase
C (PLC). The 5HT3 class of receptors are ion
channels.Some serotonin receptors are presynaptic and others postsynaptic. The
5HT2A receptors mediate platelet aggregation and smooth muscle
contraction. The 5HT2C receptors are suspected in control of food
intake as mice lacking this gene become obese from increased food intake and
are also subject to fatal seizures. The 5HT3 receptors are present
in the gastrointestinal tract and are related to vomiting. Also present in the
gastrointestinal tract are 5HT4 receptors where they function in
secretion and peristalsis. The 5HT6 and 5HT7 receptors
are distributed throughout the limbic system of the brain and the 5HT6
receptors have high affinity for antidepressant drugs. Melatonin is derived
from serotonin within the pineal gland and the retina, where the necessary
N-acetyltransferase enzyme is found. The pineal parenchymal cells secrete
melatonin into the blood and cerebrospinal fluid. Synthesis and secretion of
melatonin increases during the dark period of the day and is maintained at a
low level during daylight hours. This diurnal variation in melatonin synthesis
is brought about by norepinephrine secreted by the postganglionic sympathetic
nerves that innervate the pineal gland. The effects of norepinephrine are
exerted through interaction with b-adrenergic receptors. This leads to
increased levels of cAMP, which in turn activate the N-acetyltransferase
required for melatonin synthesis. Melatonin functions by inhibiting the
synthesis and secretion of other neurotransmitters such as dopamine and GABA.
Creatine is
synthesized in the liver by methylation of guanidoacetate using SAM as the
methyl donor. Guanidoacetate itself is formed in the kidney from the amino
acids arginine and glycine.
Synthesis of creatine and creatinine
Creatine is
used as a storage form of high energy phosphate. The phosphate of ATP is
transferred to creatine, generating creatine phosphate, through the action of creatine
phosphokinase. The reaction is reversible such that when energy demand
is high (e.g. during muscle exertion) creatine phosphate donates its phosphate
to ADP to yield ATP.Both creatine and creatine phosphate are found in muscle,
brain and blood. Creatinine is formed in muscle from creatine phosphate by a
nonenzymatic dehydration and loss of phosphate. The amount of creatinine
produced is related to muscle mass and remains remarkably constant from day to
day. Creatinine is excreted by the kidneys and the level of excretion
(creatinine clearance rate) is a measure of renal function.
Glutathione (abbreviated GSH) is a tripeptide composed of glutamate,
cysteine and glycine that has numerous important functions within cells. It
serves as a reductant, is conjugated to drugs to make them more water soluble,
is involved in amino acid transport across cell membranes (the -glutamyl cycle), is a part of the peptidoleukotrienes, serves as a cofactor for some
enzymatic reactions and as an aid in the rearrangement of protein disulfide
bonds.
Synthesis
of glutathione (GSH) Structure of GSSG
The role of GSH as a reductant is extremely important particularly in the
highly oxidizing environment of the erythrocyte. The sulfhydryl of GSH can be
used to reduce peroxides formed during oxygen transport. The resulting oxidized
form of GSH consists of two molecules disulfide bonded together (abbreviated
GSSG).
The enzyme glutathione reductase
utilizes NADPH as a cofactor to reduce GSSG back to two moles of GSH. Hence,
the pentose phosphate pathway is an extremely important
pathway of erythrocytes for the continuing production
of the NADPH needed by glutathione reductase. In fact as much as
10% of glucose consumption, by erythrocytes, may be mediated by the pentose
phosphate pathway. Several mechanisms exist for the transport of amino acids
across cell membranes. Many are symport or antiport mechanisms that couple
amino acid transport to sodium transport. The -glutamyl cycle is an example of a group transfer mechanism of amino acid transport.
Although this mechanism requires more energy input, it is rapid and has a high
capacity. The cycle functions primarily in the kidney, particularly renal
epithelial cells. The enzyme -glutamyl transpeptidase is
located in the cell membrane and shuttles GSH to the cell surface to interact
with an amino acid. Reaction with an amino acid liberates cysteinylglycine and
generates a -glutamyl-amino acid which is transported into the
cell and hydrolyzed to release the amino acid. Glutamate is released as
5-oxoproline and the cysteinylglycine is cleaved to its component amino acids.
Regeneration of GSH requires an ATP-dependent conversion of 5-oxoproline to
glutamate and then the 2 additional moles of ATP that are required during the
normal generation of GSH.
.
One of the earliest signals that cells have entered
their replication cycle is the appearance of elevated levels of mRNA for ornithine
decarboxylase (ODC), and then increased levels of the enzyme, which is
the first enzyme in the pathway to synthesis of the polyamines. Because of the
latter, and because the polyamines are highly cationic and tend to bind nucleic
acids with high affinity, it is believed that the polyamines are important
participants in DNA synthesis, or in the regulation of that process.
The key features of the pathway are that it involves putrescine, an ornithine catabolite, and
S-adenosylmethionine (SAM) as a donor of 2 propylamine residues. The first
propylamine conjugation yields spermidine
and addition of another to spermidine yields spermine.The
function of ODC is to produce the 4-carbon saturated diamine, putrescine. At
the same time, SAM decarboxylase cleaves the SAM carboxyl
residue, producing decarboxylated SAM (S-adenosymethylthiopropylamine), which
retains the methyl group usually involved in SAM methyltransferase
activity. SAM decarboxylase activity is regulated by product
inhibition and allosterically stimulated by putrescine. Spermidine
synthase catalyzes the condensation reaction, producing spermidine and
5'-methylthioadenosine. A second propylamine residue is added to spermidine
producing spermine.The signal for regulating ODC activity is unknown, but since
the product of its activity, putrescine, regulates SAM decarboxylase
activity, it appears that polyamine production is principally regulated by ODC
concentration.The butylamino group of spermidine is used in a posttranslational
modification reaction important to the process of translation. A specific lysine residue in the translational initiation factor eIF-4D is modified. Following the modification the
residue is hydroxylated yielding a residue in the protein termed hypusine.
Nitric Oxide
Synthesis and Function
Vasodilators, such as acetylcholine, do not exert their effects upon the
vascular smooth muscle cell in the absence of the overlying endothelium. When
acetylcholine binds its receptor on the surface of endothelial cells, a signal
cascade, coupled to the activation phospholipase C- (PLC), is initiated. The
PLCg-mediated release of inositol trisphosphate, IP3 (from membrane
associated phosphatidylinositol-4,5-bisphosphate, PIP2), leads to
the release of intracellular stores of Ca2+. In turn, the elevation
in Ca2+ leads to the liberation of endothelium-derived
relaxing factor (EDRF) which then diffuses into the adjacent smooth
muscle. Within smooth muscle cells, EDRF reacts with the heme moiety of a
soluble guanylyl cyclase, resulting in activation of the latter
and a consequent elevation of intracellular levels of cGMP. The net effect is
the activation of cGMP-responsive enzymes which lead to smooth muscle cell
relaxation. The coronary artery vasodilator, nitroglycerin, acts to increase
intracellular release of EDRF and thus of cGMP. Quite unexpectedly, EDRF was
found to be the free radical diatomic gas, nitric
oxide, NO. NO is formed by the action of NO synthase,
(NOS) on the amino acid arginine.
Nitric oxide is involved in a number of other important cellular processes
in addition to its impact on vascular smooth muscle cells. Events initiated by
NO that are important for blood coagulation include inhibition of platelet aggregation and
adhesion and inhibition of neutrophil adhesion to platelets and to the vascular
endothelium. NO is also generated by cells of the immune system and as such is
involved in non-specific host defense mechanisms and macrophage-mediated
killing. NO also inhibits the proliferation of tumor cells and microorganisms.
Additional cellular responses to NO include induction of apoptosis (programmed
cell death), DNA breakage and mutation. NOS is a very complex enzyme, employing
five redox cofactors: NADPH, FAD, FMN, heme and tetrahydrobiopterin. NO can
also be formed from nitrite, derived from vasodilators such as glycerin
trinitrate during their metabolism. The half-life of NO is extremely short,
lasting only 2-4 seconds. This is because it is a highly reactive free radical
and interacts with oxygen and superoxide. NO is inhibited by hemoglobin and
other heme proteins which bind it tightly.Chemical inhibitors of NOS are
available and can markedly decrease production of NO. The effect is a dramatic
increase in blood pressure due to vasoconstriction. Another important
cardiovascular effect of NO is exerted through the production of cGMP, which
acts to inhibit platelet aggregation.
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