Amino Acid Synthesis Flashcards
What is the source of nitrogen for amino acid synthesis?
Atmospheric Nitrogen (N2)
- abundant- 80%
- Triple Bond is very strong-extrememly unreactive
Sources of Nitrogen Fixation
1) 60% Diazotrophic (nitrogen fixing) microorganisms
- Rhizobium Bacteria
2) 15% lighting and UV light
3) 25% commercial process
- habers process
- N2 + 3H2-> 2NH3
- requires 300 atm, 500 Degrees F, Iron Catalyst
Nitrogen Fixation by Rhizobium Bacteria
-Nitrogenase complex
Nitrogenase complex
1) reductase
- Provides 8e- from reduced ferredoxin with high reducing potential
- ATP hydrolysis used to transfer e- to nitrogenase (2ATP/ e-)
2) Nitrogenase
- uses electrons provided by reductase to reduce N2 to NH3+
Rhizobium Bacteria: Reductase structure
AKA
- iron proteins
- Fe protein
Dimer of identical subunits
1) 4Fe-4S cluster
- bridges subunits
- transfers E- ONE AT A TIME to Nitrogenase
2) each subunit contains ATP Binding domain (Ploop)
Rhizobium Bacteria: Nitrogenase Structure
AKA:
- Molybdenum-iron protein
- MoFe protein
A2B2 Tetramer 1)FeMe Cofactor -uses electrons from P cluster to reduce N2 to NH3 -2 M-3Fe-3S custers M=Mo in one cluter M= Fe in other cluster *organization unique to nitrogenase 2) P cluster -stores e- before transfer and use by FeMo
This ENYZME STILL A SUBJECT OF ACTIVE RESEARCH
Source Of Component for Amino Acid Synthesis?
Nitrogen:
-Glutamate from Ammonia
Carbon Skeleton:
- Pyruvate
- Acetoacetyl CoA
- Acetyl CoA
- OAA
- Fumurate
- Succinyl CoA
- A-ketogluterate
Humans can synthesize the 11 nonessential amino acids. What are they?
Alanine, Arginine, Asparagine, Aspartate
Glutamate, Glycine, Glutamine
Cysteine, Serine, Tyrosine
Proline
Glutamate Dehydrogenase
Assimilates Ammonium Ion (NH4+) into Glutamate (two step process)
-Transamination of alpha amino group nitrogen of glutamate provides (most) amino acids with alpha amino group
Mechanism:
1) Schiff Base between ammonia and carbonyl of a-ketogluterate
- replace C=O with C=NH2+, and releases H2O
2) Protonated Schiff Base is reduced by transfer of hydride from NADPH
Estabilishes stereochemistry
Common Feature of amino acid synthesis
Formation of Schiff Base followed by protonation and reduction
Synthesis of Glutamine
Glutamine Synthetase
Amidation of R group of glutamate produces glutamine
Precursor-Glutamate
Activated by:Phosphorylation-Phosphate added to R group carboxylic acid
-Phosphate is displaced by NH3+ to form Glutamine
Synthesis of Aspartate
Aspartate Transaminase
-synthesized from a-ketoacids in one step by PYRIDOXAL PHOSPHATE-dependent transaminases
Precursor-OAA
OAA + Glutamate -> Aspartate + a-ketogluterate
Synthesis of Alanine
Alanine Transaminase
-synthesizes from a-ketoacids in one step by Pyridoxal Phosphate-dependent transmaminases
Precursor: Pyruvate
Pyruvate + Glutamate -> Alanine + a-ketogluterate
Synthesis of Asparagine
Asparagine Synthetase
Precursor: Aspartate
Activation- Adenylation
-Asparate adenylated to form Acyl Adenylate intermediate
-Glutamine provides NH3+ which displaces AMP to form Asparagine
Synthesis of Proline
Multiple Steps
Precursor: Glutamate
Activated- Phosphorylation
-Acyl-Phosphate Int is reduced to Glutamic Y-semialdehyde at the expense of NADPH to NADP+
-Glutamic Y-Semialdehyde dehydrated(No enzymatic reaction-spontaneous) then reduced to Proline at the expense of NADPH to NADP+
Synthesis of Arginine
Multiple Steps
Precursor: Glutamate
Activated- Phosphorylation
-Acyl Phosphorylate Intermediate is reduced to Glutamic y-semialdehyde at the expense of NADPH to NADP+\
-Glutamate transfers amino group to Glutamic Y-semialdehyde to form Ornithine
-Ornithine enters Urea Cycle-> Arginine
Synthesis of Serine
Multiple steps
Precursor: 3-Phosphoglycerate
-3-PG oxidized to 3-phosphohydroxypyruvate at the expense of NAD+ to NADH
-Glutamate transfers Amino group to 3-phosphohydroxylpyruvate forming a-ketogluterate + 3-Phosphoserine
-3-phosphoserine is hydrolyzed displacing the Phosphate with OH to produce Serine
Synthesis of Glycine
2 Pathways
1)Serine hydroxymethyl transferase
-reversing this enzyme can be used to produce serine from glycine
-allows one carbon units to be synthesized from carbohydrates
Precursor- Serine
Serine + THF -> glycine + methylenetetrahydrofolate + H2O
2)Serine Synthase (glycine cleavage enzyme)
Precursor: NH4 + CO2
NH4 + CO2 + N5,N10methyleneTHF + NADH-> GLYCINE + THF + NAD+
THF
tetrahydrofolate=COFACTOR
Carries activated one carbon units
-one carbon units attach to N5 or N10 or both
-one carbon units are interconvertible while attached to THF
Humans unable to synthesize
-obtained from diet or intestinal microorganisms
SAM
S-Adenosylmethionine
Methyl Group donor as part of activated methyl Cycle
- Synthesized from Met and ATP
- Methyl group is activated due to + charge on sulfur
ALL 3 phosphates are lost
-hydrolyzed into Pi (orthophosphate) and PPi (pyrophosphate)-> further hydrolyzed to 2Pi
Activated Methyl Cycle
Regenerates Methionine from homocysteine
1) Methionine + ATP-> S-adenoysl Methionine (SAM)
2) SAM releases Activated CH3 to form S-adenosyl Homocysteine
3) S-Adenosyl Homocysteine is hydrolyzed to form homocysteine+ adenosine
4) Homocysteine is methylated (-CH3)-> Methionine
- catalyzed by Methionine synthase (aka homocysteine methyl transferase)
- N5-methyl THF-> THF
- requires Methylcobalamin derived from Vit B12
Synthesis of Cysteine
2 steps:
Precursor: Serine and Homocysteine
1)Cystathionine B-Synthase
Homocysteine + Serine-> Cystathionine + H2O
-Ser R group attaches to Sulfur and loses O
2)Cystathioninase
Cystathionine hydrolyzed to NH4+ + a-ketobutyrate + CYSTEINE
Synthesis of Tyrosine
Phenylalanine Hydroxylase (monooxygenase)
Precursor: Phenylalanine
Phe + O2+ tetrahydrobiopterin-> Tyr + H2O + quinoid dihydrobiopterin
-Hydroxylates
-in humans Phe is essential if Phe is not present then tyrosine becomes essential
Synthesis of Amino Acids that are essential in humans are found in?
Plants and bacteria
Plant and Bacteria:
-Phenylalanine, tryptophan, Tyrosine synthesis
Bacteria: substrates
- Phophoenolpyruvate from glycolysis
- erythrose 4-Phosphate from Pentose phosphate pathway
Intermediates
- Shikimate after phosphorylation can be inhibited by Glyphosate (round up)
- Chorismate-common intermediate for aromatic aa **
Substrate channeling
Indole is hydrophobic and would diffuse out of cell thus use Substrate channeling:
- Substrate goes through channel and is passed from active site to active site until released
- speeds up rate of synthesis
Chirality of Amino Acids
-all aminotransferases contain 2 conserved amino acids?
Lysine that forms the Schiff base with PLP
-Arginine that interacts with alpha-carboxylate group of the ketoacid
Methionine Synthase
AKA homocysteine methyltransferase
- requires methylCobalamin derived from Vit B12
- this enzyme exists in some organisms that doesn’t require methylcobalamin
Homocysteine is methylated by N5,methyl THF-> methionine + THF
PRPP
activated form of ribose
Negative Nitrogen Balance
Deficiency of even one amino acid leads to this
More proteins degraded then synthesized, and more nitrogen is excreted then ingested
What do all aminotransferases contain?
- Lysine that forms Schiff base with PLP
- Arginine that interacts with alpha carboxylate group of the ketoacid
Glutathione
Sulfhydryl Buffer and Antioxidant that protects us from Reactive Oxygen Species (ROS)
Structure- Tripeptide w/Sulfhydryl ECG
-Glutamate R group attaches to Cysteins N displace O-
GSH-> GSSG
Glutathione reductase
-Uses NADPH to reduce GSSG to GSH
-Prosthetic group-FAD
Glutathione Peroxidase
Reduces hydrogen peroxide to water
Selenium (Se) replaces Sulfur in R group of cysteine active site
-E-Se-(selenolate) reduces peroxide to hydroxyl while being oxidized to to an acid E-S-OH (Seleninic acid)
-E-S-OH is then reduced by GSH which is oxidized to GSSG
-GSSG then reduced by NADPH to NADP+
Nitric Oxide
Second messenger in signal transduction pathway
- synthesized from Arginine, NADPH, and O2
- enzyme is another reason why animals must inhale O2
Nitric Oxide Synthase
1) Arginine ->N-w-hydroxyarginine
- O2 (monooxygenase)-> H2O
- NADPH to NADP+
2) N-w-Hydroxyarginine-> Citrulline + NO
- O2-> H2O (monooxygenase)
- NADPH- to NADP+
Porphyrins
-no mechanism just general info
used to synthesize heme
- synthesized from Succinyl CoA and glycine
- cyclic compounds that readily bind iron
Where does Heme Synthesis occur?
-Compartmentalization?
Cellular Localization
- Mitochondria
- Cytoplasm
Organ Localization
- Liver=variable rate
- Erythrocytes producing cells of bone marrow: constant rate
S-aminolevulinate Synthase
1) S-aminolevulinate Synthase
Gly + Succinyl CoA -> S-aminolevulinate (ALA or S-aminlevulinic acid)
-Amino Group of Glycine attaches to Carbonyl of Succinyl CoA
-releases CoA and CO2
-occurs in matrix of mitochondria
8 S-aminoleveulinate is required to synthesize one heme
S-aminolevulinic dehydratase
S-aminolevulinic (x8)-> (x4) Porphobilinogen
- occurs in cytosol
- two molecule are joined to form porphobilinogen
*Lead inhibits causing the anemia associated with lead poisoning
Hydroxymethylbilane synthase
-links 4 porphobilinogen molecules to form linear tetrapyrole
What are the last 3 steps in Porphyrin synthesis
Uroporphyrinogen III synthase
- cytosol
- cyclized linear tetra pyrrole to form ring
Uroporphyrinogen carboxylase
- cytosol
- produce coproporphyrinogen III has methyl instead of A group
Coproporphyrinogen-> Protoporphyrin IX
- matrix of mitochondria via multiple steps
- Fe added is final step to form HEME
Ferritin
Iron storage protein
Transferrin
Iron transport protein
Ferrochelatase
enhances rate of addition of iron (Fe2+) to protoporphyrin IX
-this may occur without enzyme as well
Degradation of Heme
1) Heme-> Biliverdin
- O2-> H2O (monooxygnease)
- releases Fe3+
- NADPH-> NADP+
2) Biliverdin-> Bilirubin
- Biliverdin reductase
- NADPH to NADP+
