Transcript AA lecture 2 urea cycle

Section 8.
Amino Acid
Metabolism
Urea cycle
11/18/05
Substrates for the Urea Cycle
-amino acid
-keto acid
-ketoglutarate
glutamate
NADH  
H2N-CO-OPO32-
NAD+ 
• Above, amino groups are transferred to glutamate,
from which ammonium is produced, and then used to
make carbamoyl phosphate.
• Below, amino groups are transferred to produce
aspartate.
-amino acid
-ketoglutarate
-keto acid
glutamate
1
aspartate
oxaloacetate
Urea Cycle
Fig. 23.16
2
• Aspartate and
carbamoyl
phosphate
each deliver an
amino group to
the cycle.
• Notice that the
carbamoyl
phosphate
production and
condensation
occur in the
mitochondrial
matrix.
NH4+ from Oxidative Deamination of Glutamate
O O
+
NH3
H
CH2
CH2
O
O
glutamate
NAD+
NADH + H+
O
O
O
CH2
CH2
glutamate
dehydrogenase
O
+ NH4+
O
-ketoglutarate
• Hexameric glutamate dehydrogenase is is controlled
allosterically.
– High energy levels inhibit (ATP and GTP).
– Low energy levels activate (ADP and GDP).
• NADP+ can replace NAD+.
• NH4+ , which is toxic to humans, is produced in the
mitochondria and used to make carbamoyl phosphate.
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Carbamoyl Phosphate Synthesis
(p. 645)
• Carbamoyl phosphate synthetase is in mitochondrial matrix.
• NH4+ is source of NH3.
• The hydrolysis of two ATP make this reaction essentially
irreversible.
4 • N-acetyl glutamate is an allosteric activator. (see S08L05)
O
H
O
N
CH 2
CH 2
Pi +
O
O P O
O
H2N
CH 2
H
carbamoyl
phosphate
O
O
H
+
H 3N
CH 2
+ ATP
+
NH 3
O
O
aspartate
O
O
4
NH 2
citrulline
1
+
NH 3
+
CH 2
ornithine
UREA
CYCLE
CH 2
CH 2
+
H
NH 3
O
2 ~ P used
AMP + PPi
2 Pi
+
H
NH 3
O
O
+
3
H2N
O
O
H
N
N
+
H CH 2
CH 2
CH 2
O
O
CH 2
H
(1-3 in cytosol, 4
in mitochondial matrix)
O
O
H 2N
NH2
UREA
H2O
H 2N
H
N
CH 2
CH 2
NH 2
CH 2
+
H
NH 3
O
O
argininosuccinate
2
O
H
O
H
O
O
fumarate
arginine
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1. ARGININOSUCCINATE SYNTHASE 2. ARGININOSUCCINASE
3. ARGINASE
4. ORNITHINE TRANSCARBAMOYLASE
Connection to Krebs Cycle
+ ATP
-keto acid
aspartate
citrulline
-amino acid
1
UREA
CYCLE
Oxaloacetate
argininosuccinate
+ AMP + PPi
NADH
2 Pi
NAD+
2
arginine
Malate
fumarate
H20
6
• Fumarate is oxidized to oxaloacetate by Krebs cycle
enzymes, producing NADH.
• Oxaloacetate accepts an amino group instead of
being condensed with acetyl CoA.
amino
acid
-keto
acid
-ketoglutate
glutamate
*
NH4+ + CO
2
Amino Acids to Urea
+ 2 ATP + H2O
2 ADP + Pi + H+ + carbamoyl
phosphate
ornithine
+ ATP
citrulline
UREA
CYCLE
-keto acid
aspartate
amino
acid
argininosuccinate
Oxaloacetate
+ AMP + PPi
H2N
O
UREA
2 Pi
NH2
Malate
arginine
H2O
fumarate
*Glutamate Dehydrogenase is the control site: ADP (+),
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GDP (+), ATP (-), GTP (-) and NADH (-).
Control at other sites by glucagon (+), cortisol (+), insulin
(-), growth hormone (-).
Summary of Reactions and Energetics - 1
H20 + aa + NAD+  -keto acid + NH4+ + NADH + H+
and
H20 + fumarate + aa + NAD+  aspartate + -keto acid
+ NADH + H+
then
aspartate + NH4+ + HCO3- + 3 ATP 
urea + fumarate + 2 H20 + 2 ADP + AMP + 4 Pi + H+
Four high energy phosphate bond equivalents are
used for these reactions (- 4 ~P).
Two NADH are produced.
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Summary of Reactions and Energetics - 2
Now consider NADH oxidation:
2 H+ + 2 NADH + O2 2 NAD+ + 2 H20
(+5 ~P)
The net reaction is then
2 aa + HCO3- + O2 
2 -keto acid + urea + H+ + 2 H20
(+1~P)
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Hyperammonemia
• Normal blood [NH4+] is 10-40 mM.
• Deficiencies of carbamoyl phosphate synthetase or
of any enzyme in the urea cycle cause high [NH4+].
• Affects CNS and can lead to irreversible brain
damage.
• Treatment strategies depend on which enzyme is
deficient.
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• Low dietary protein
reduces need for urea
cycle.
H
O
NH 3
• High dietary arginine
H
+ ATP provides a path for
CH
carbamoyl phosphate
O
O
and aspartate
aspartate nitrogens to produce
argininosuccinate,
O O
which is excreted.
HN
CH 2
H
O
O
O
O P
O
H2N
Argininosuccinase Deficiency
N
CH 2
CH 2
NH 2
H 3N
H
4
+
NH 3
O
O
citrulline
1
+
ornithine
+
2
UREA
CYCLE
CH 2
CH 2
+
H
NH 3
O
+
O
H2N
NH2
UREA
H2O
H
N
CH2
CH2
11
+
AMP + PPi
2 Pi
+
NH 3
O
O
NH2
CH2
+
H
NH3
O
H
N
N
H CH 2
CH 2
CH 2
O
O
CH 2
H
H2N
3
O
O
2
CH 2
carbamoyl
phosphate
O
+
argininosuccinate
2
(excreted)
O
H
O
H
O
O
fumarate
O
arginine (excess supplied)
Carbamoyl Phosphate Synthetase Deficiency
Fig. 23.20
12
• Hippurate and phenylacetylglutamine are excreted.
• Amino groups to glycine and glutamine by transamination.
Ketogenic and Glucogenic Amino Acids
Fig. 23.21
• After removal of the amino group, the keto acids are used to make
13 Krebs cycle intermediates, pyruvate, acetyl CoA and acetoacetyl CoA.
Nitrogen for Oral Bacteria
H2NCONH2 + H+ + H2O
urease
2 NH4+ + HCO3-
• Urea is a major source of nitrogen for oral
bacteria.
• It diffuses through most membranes and is
in saliva.
• Bacterial urease produces NH4+.
• Glutamate dehydrogenase incorporates
NH4+ into -keto acids to obtain amino
acids for bacterial growth.
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Nitrogen for Bacterial Amino Acid Synthesis
NADPH + H+
-ketoglutarate
+ NH4+
glutamate
glutamate
dehydrogenase
ATP
glutamate
+ NH4+
NADP+
+ H20
ADP + Pi
glutamine
glutamine
synthase
NADPH + H+
-ketoglutarate
+ glutamine
NADP+
2 glutamate
glutamate
synthase
• When [NH4+] is limiting, it does not bind glutamate dehydrogenase,
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and the lower two reactions are used.
Engineered Oral Bacteria to Fight Caries?
• Streptococcus Salivarius urease activity affects
oral microbial ecology.
• It produces NH3, which in addition to promoting
growth, neutralizes acids produces by other
bacteria.
• S. Salivarius urease gene was introduced into
Streptococcus mutans GS5. It was expressed
and during glucose metabolism reduced pH
decrease and duration.
•
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(Clancy & Burne,1997 FEMS Microbiol Lett 151:205)
Web links
Nitrogen Fixation. A summary of the topic.
Nitrogen Cycle. The biological big picture.
Amino Acid Metabolism. Reviews reactions.
Next topic: Porphyrins, heme,
bile pigments