Dr. Purich - Biochem Flashcards
What is the purpose of glutamine synthase?
It is responsible for the production of glutamine in the cytosol.
How is the glutamine sythase reaction powered?
We use ATP to power the trapping of ammonia to keep ammonia levels low and synthesize glutamine.
How does GDP use different cofactors for opposite reactions?
The overall reactions are:
NAD+ + Glu –> Alph-KG + NH4 + NADH
NADP+ + Glu –> Alpha-KG + NH4 + NADPH
The ratio of (NAD/NADH) = 700
The ration of (NADP+/NADPH) = .0005
This drives the first reaction to the right and second to the left. This means that the very same enzyme in the presence of different cofactors catalyzes to different reactions.
Where is CPS2 found?
It is found in the cytosol and is used in the production of pyrimidine nucleotides. It uses glutamine as a substrate.
What is the amino acid acceptor for transamination reaction to proceed?
alpha-ketoglutarate
If an alpha-ketogluterate has an amino acid bound to it how do we remove it?
By deamination. This regenerates the alpha-ketogluterate and also forms nitrogen.
When NAGS is activated what happens?
The cell makes N-Acetyl-Glutamate from Glutamate and acetyl-Coa
What are some of the characteristics of the urea pathway?
It is highly energy dependent
It releases considerable acid in the process
What factors dictate protein turnover?
- Turnover rate is encoded in protein
- Metabolism of person
If you have high Glutamate and no good NAGS what happens?
The cell can’t produce Urea from the excess Glutamate and becomes hyperammonemic.
What is the main nitrogenous product in mammals?
Urea - it is the end product of ammonia detoxification.
For what reasons does de novo amino acid synthesis occur?
- To provide building blocks for protein
- Restore amino acid pools in specific tissue
- Needed for synthesizing special AA’s
- As a source of energy
- Glucogenic (make glucose)
- Ketogenic (make ketones)
How do we get single amino acids and very small peptides?
By the actions of other pepsidases (exo in epithelial and carboxyl is soluble)
When does cys become an essentail amino acid?
Then there are low levels of Met
Once you have Argininosuccinate in the urea cycle what happens next?
We split it up with a lyase - argininosuccinate lyase and produce arginine and fumarate. Clinical manifestations of excess urea can be minimized by decreasing protein intake and reducing starvation, steroid use.
What kinds of NAGS mutations are there?
Primary: Mutation in gene
Secondary: Mitochondrial changes that interfer with NAGS
What is a non-essential amino acid?
One that can be manufactured in the body de novo
Examples of transamination reactions that have a Keq=0 are:
Alanine + alph-ketoglutarate = pyruvate and glutamate
Glutamate + Oxaloacetate = Aspartate + α-Ketoglutarate
What enzye is responsible for deamination glutamate?
Glutamate Dehydrogenase
What are the two major pathways of protein turnover?
- Ubiquitin dependent pathways that use proteosomes
- Lysosomal / phagocytic pathways that break them apart after H+ has made them neutral and set them up for proteolysis.
What organ is responsible for the processing of excess ammonia into urea?
The liver
What is the primary nitrogenous end product excreted by mammals?
Urea
What dictates the amount of urea we excrete?
Metabolism rate
What is the primary point of the urea cycle?
To detoxify ammonia.
What NAGS analogue can be used to restore glutamate degradation?
N-Carbamoylglutamate
The urea cycle happens where?
In both the cytosol and mitochondria of liver cells.
What is an essential amino acid?
What that must be taken up in the diet - the body can’t make it de novo
What is common to amino acids derived from diet and thos that are released from degredation pathways?
The all enter the same pathways of biosythesis.
What do L and D amino acid oxidases do?
They take an amino acid and oxygen in the presence of FMN and forms NH3, alpha Keto Acid and H2O2. This is a reduction of O2 to H2O2.
What multiple roles does glutamine play?
- Very good at trapping excess NH3 keeping it from dangerous circulation.
- Provides the nitrogen for many biosynthetic reactions.
- Functions as a nitrogen shuttle between organs.
Where are proteins first fragmented and how?
In the stomach by pepsin (pH 1-2)
In the duodenum by trypsin and chymotrypsin (pH 7)
This results in small peptides
What happens to glutamine as it travels through the liver acinus from portal to central end?
It starts off with high amounts of gluatmine and is processed by glutaminase and the urea cycle to make urea. This process mops up most of the NH3 - urea sythesis has a high Km (low affinity for NH3)
Toward the central end of the acinus any remaining NH3 that is left can get captured by the glutamine synthase which has a high affinity for NH3 (low Km) and make sure no excess NH3 enters circulation.
What is the end result of pepsin, trypsin, chymotripsin, carboxylpepsidase and exopepsidase?
Small single amino acids and peptides that are absorbed in the intestines. Often by a Na+ gradient or energy source.
How is nitrogen being excreted by the body?
- Ammonia
- Uric Acid
- Urea
When deamination is done in the liver using glutaminase what are the products?
Ammonia (goes into Urea cycle)
Glutamic acid
What is positive nitrogen balance and why do we need it?
It is when we intake more nitrogen than we excrete. We need a positive nitrogen balance help with growth, wound healing, pregnancy and recovering adults.
What is one way that Amino Acids exit the intestinal lumen and enter the portal vein?
By way of a sodium powered symporter. The gradient is created by the Na/K ATPsynthase. This is the same way that the Na / Glucose symporter works.
How does the nitrogen created by GDH get used?
Only in the urea cycle.
What does aspargine and h2o for in the presence of asparaginase?
This is a different elimination reaction in which aspartate and ammonia form. This is hydrolytic deamination.
Once you have citrulline what happens next in the urea cycle and by what enzyme?
The cell forms Argininosuccinate using argininisuccinate sythetase. This is ATP dependent. If this enzyme is broken we have citrullinemia 1 - hyperammonia, increased citrulline, seizures, loss of consciousness and death.
What does high ammonia do in the cell?
Competes with other amine compounds in certain certain enzyme catalyzed reactions.
Crosses membranes and wreck ion transport mechanisms - leads to alkalosis.
What does GDH use when doing oxidation deamination?
NAD+
What controls the Urea cycle?
High protein - increases urea cycle
Glucagon - increases urea cycle
Some amino acids - increase glucagon –> increase urea cycle
Arginine –> increases urea cycle
Lots of AA –> glutamate –> N-acetylglutamate (arginine stimulated)
Once you have arginine from the urea cycle what happens next.
The cell makes ornithine and UREA using arginase as the enzyme. The best way to diagnose and arginase deficiency is 3-4x normal levels in the plasma.
When is Arg an essentail amino acid?
When the rate of growth in children and pregnant women is too fast for de novo synthesis.
Where is CPS1 found?
In the mitochondria and is responsible for binding NH3 and formation of carbomyl-phosphate using 2 ATP.
Enzyme bound PLP pyridoxal-phosphate is converted to?
Enzyme bound PMP pyridoxamine-phosphate. This is the process of transamination.
What liver cell is responsible for the processing of ammonia into urea?
The liver cell acinus.
Are animal based diets always sufficient in essentail amino acids?
No, low amounts of methionine, arginine and lysine.
What doe a B6 deficiency affect?
Many amino acid driven reations as well as the formation of neurotransmitters that are derivatives of amino acids.
What allows the liberation of NH3 and NADH in the mitochondria?
The combination of a transaminase and a GDH we can liberate ammonia and NADH in the mitochondria. This is accomplished by the nonspecific reaction of a transaminase.
What does OTC (Ornithine Transcarbamoylase) do?
It is an enzyme in the Urea cycle that transferes a carbamyl group from carbamyol-p to orthinine to form citruline.
OTC deficiency is x-linked inherited in male neonates and results in hyperammonemic encephalopathy.
If you see Gout in your clinic what nitrogenous excretion product is this from?
Uric Acid
Where does glutamate dehydrogenase (GDH) work?
In the mitochondrial matrix. ** This also creates an NADH in the mitochondrial matrix**
What happens to histidine in the presence of histidinase?
This is also a deamination but we also create a C-C double bond so it is elimination deamination. We still get ammonia. A buildup of this can cause histidinemia.
What are the sources of Amino Acids?
Dietary: Both essentail and nonsessential
De Novo amino acid synthesis
Protein turnover in the body
Glutamate + Acetyl-CoA =
N-Acetyl-Glutamate and CoA
When does Tyr become and essentail amino acid?
At low levels of Phe
What are some of the general principles of amino acid synthesis and nitrogen metabolism?
- Humans are the only mammals that chose their amino acid intake.
- All mammals take in some amino acids in greater quantity than they need.
- Amino acid metabolism is part of nitrogen metabolism.
- Amino acids that are not used in proteins are not re-excreted. This is why amino acids in urine is indicator for illness
- Excess AA are used for their carbon skeleton to make energy or storage as fat.
What is true nitrogen balance?
The intake and excretion of nitrogen is equal
Where are the amino acids ultimately delivered?
- Pyruvate –> alinine
- α-ketoglutarate –>glutamate
These are both intermediates of the TCA.
Transamination reactions are fully reversible why?
Because the delta-G is nearly 0 and the reactions proceed forward and backward almost equally. This makes Keq=0.
What population has higher amino acid needs?
Children and pregnant women
What is another way we can deaminate amino acids?
We can use hydrolytic deamination using glutaminase. This is less active than using glutamate dehydrogenase but is done nonethless.
How is Carbamoyl-Phosphate Synthetase important in the urea cycle?
It is the enzyme that catalyzes the first committed step of the urea cycle.
Transamination requires a cofactor. What is it and from what is it derived?
pyridoxal phosphate derived from vitamin B6. What would be the result of a vitamin B6 deficiency?
What enzyme is activated by high levels of arg and glu?
NAGS (N-Acytl-Glutamate Synthase)
What four amino acids are not transaminated?
Proline
Hydroxyproline
Lysine
Threonine
What is negative nitrogen balance and its affects?
When we excrete more nitrogen than we take in. The results:
Starvation
Malnutrition
Disease
Why is carbomyl phosphate important?
It is a powerful carbomylating agent and is left attached to attacking nucleophiles that displace a phoshpate group.
What is Carbamoyl-Phoshpate?
The formation of carbomyl-phosphate is the first committed step in shuttling NH3 into the urea cycle. It requires two ATP and has a two step enzyme bound process with intermediates. The enzyme that catalyzes the reaction is carbomyl-phosphate synthase.
What are the essentail amino acids?
PVT TIM HLL
Phen
Valine
Trp
Thr
Ile
Met
His
Leu
Lys
Arginine is nonessential how is it made?
Manufactured via the conversion of citrulline to argininosuccinate via ASS and arginine and fumarate via ASL.
Glutamate is a nonessentail amino acid, how is it manufactured?
Via four possible reactions.
- Transamination
- GDH
- Glutamate synthase from glutamine
- Glutaminase from glutamine
Apartate is a nonessential amino acid, how is it manufactured?
Via transamination of OAA and glutamate. This is the energetically equivalent reaction to Alanine –> pyruvate via alpha-ketogluterate.
It can also be hydrolysised via apsaraginase.
Asparagine (Asn) is a nonessential amino acid how is it made de novo?
Asparagine (Asn) is made via aspargine synthase from aspartic acid + glutamine +ATP.
