Nitrogen Metabolism Nitrogen Forms in the Body Nitrogen Balance Critical Body Must Make and Break Down Amino Acids Nitrogen Also Needed for Synthesis of Nucleotides ATP GTP CTP UTP dATP dCTP dGTP dTTP ID: 659770
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Slide1
Nitrogen Metabolism
Dr. Kevin AhernSlide2
Nitrogen MetabolismSlide3
Nitrogen Forms in the Body
Nitrogen Balance Critical
Body Must Make and Break Down Amino Acids
Nitrogen Also Needed for Synthesis of
Nucleotides (ATP, GTP, CTP, UTP, dATP, dCTP, dGTP, dTTP)
Non-Protein Amino AcidsOrnithineCitrullineSarcosineOther Nitrogen-Containing CompoundsCholineVitaminsCarnitine
Sarcosine
Citrulline
OrnithineSlide4
Excretion of Nitrogen
Amino Acids Through Transamination Make Nitrogen Mobile
Toxicity of Ammonia means Nitrogen Balance is Critical in the Body
Excretion
Ammonotelic - Excrete Ammonia - Fish
Uricotelic - Excrete Uric Acid - BirdsUreotelic - Excrete Urea - Most Vertebrates, Some Invertebrates
Ammonia
Uric Acid
Urea
Produced by Amino
Acid Catabolism
Used in Urea Cycle
Produced by Purine
Catabolism
Produced by
Urea CycleSlide5
α-ketoglutarate Family
Transamination to Make Glutamate
α-ketoacid #1
Amino Acid #2
α-keto Acid #1 +
Amino Acid X
Amino Acid #1
+ α-keto Acid X
Amino Acid X
α-ketoacid XSlide6
Glucose-Alanine Cycle
Alanine Metabolism
Glucose-Alanine Cycle
Important for Removing Ammonia
High Ammonia
Low Ammonia
Transamination in Liver
Creates Glutamate
Alanine Carries Amine to Liver
Breakdown of Glutamate
Yields Amine for Urea ProductionSlide7Slide8
Urea Cycle
Primarily Occurs in Liver. Also in Kidney
Consists of 4 Cycle Reactions and 1 Feeder Reaction
Feeder Reaction Incorporates 1 Molecule of Ammonia and 1 CO
2
Per TurnCycle Reaction Provides 1 Amine from an Amino AcidOutput of Cycle is 1 Molecule of Urea Per TurnThe Net Reaction Per Turn of the Cycle is 2 NH3 + CO2 + 3 ATP + H2O → urea + 2 ADP + 4 Pi + AMPSlide9
Urea CycleSlide10
Carbamoyl Phosphate Synthetase Reaction
2 ATP + HCO
3
−
+ NH
4+ <=> 2 ADP + Carbamoyl phosphate + PThe Source of Ammonium Ion is Glutamine or GlutamateRequires Action of Glutaminase (Glutamine) or Glutamate Dehydrogenase (Glutamate)
Glutamine + H
2O
Glutamate + NH3
Glutamate + H
2
O + NADP
+
α-ketoglutarate + NH
3
+ NADH + H
+
H
2
O + CO
2
HCO
3
-
+ NH
4
+
H
2
O + CO
2
HCO
3
-
+ NH
4
+Slide11
Ornithine Transcarbamoylase Reaction
+
Carbamoyl Phosphate
Ornithine
+
P
i
Citrulline
Phosphate
Ornithine Transcarbamoylase
Enzyme Expressed Only in Liver
Most Commonly Deficient Enzyme in Urea Cycle
X-linked Inheritance
In Severe Deficiency, Ammonia Levels Rise
to Lethal Levels if Untreated
Liver Transplant and Low Protein Diet Most
Common TreatmentsSlide12
Citrulline Transport to Cytoplasm
Citrulline Movement to Cytoplasm Requires Ornithine-Citrulline Translocase
Antiport - Moves Citrulline Out, Ornithine In
Needed for Both Parts of Urea Cycle
Deficiency of Translocase Mimics Defective Ornithine Transcarbamoylase
Condition at Birth More Serious Than Adult OnsetSlide13
Argininosuccinate Synthetase
Two Step Reaction
First, AMP Attaches to Amine-rich End of Citrulline
Next, Aspartate Displaces the AMP
The Product is L-argininosuccinate
Reaction is Rate Limiting Step of CycleGene Expression of Enzyme Reduced by Arginine, Increased by CitrullineEnzyme Defects Lead to Citrullinemia - Accumulation of AmmoniaTreated with Low Protein Diet, Arginine Supplementation
1
2Slide14
Argininosuccinate Lyase
+
Argininosuccinic Acid
Arginine
Fumaric Acid
Bond Cleaved
Important for Production of Arginine
Source of Fumarate
Deficiency Like That of Other Urea Cycle Enzymes - Ammonia Excess
Argininosuccinate Lyase
Argininosuccinate
Lyase
To Proteins or Remainder
of Urea Cycle
To Citric Acid CycleSlide15
Arginase
+
H
2
O
+
Arginine
Urea
Ornithine
Cut
Excreted
To Mitochondria
To Complete Cycle Through
Ornithine Citrulline Translocase
Arginase
Co-expressed with Nitric Oxide Synthase in Smooth Muscle
Increased Arginase Activity Reduces Nitric Oxide Production
Nitric Oxide Relaxes Smooth Muscle and Facilitates Erection of Penis
Deficiency of Arginase Rarest of Urea Cycle Enzymes
Two Forms of Arginase Provide Some Backup When One DeficientSlide16
Urea CycleSlide17
Citrulline
Alternate Means of Producing Citrulline - Nitric Oxide Synthase
Bypasses Mitochondrial Part of Urea Cycle & Produces Nitric Oxide
Arginine is Substrate for Reaction
Nitric Oxide Important Signaling Molecule in Humans - Vasodilation
2 L-arginine + 3 NADPH + 1 H+
+ 4 O2
2 Citrulline +2 Nitric Oxide + 4 H
2O + 3 NADP+
Nitric Oxide Synthase
Viagra works by enhancing signaling through the nitric oxide pathway in the penisSlide18
Nitrites
Nitrite formed by Ionization of Nitrous Acid (HNO
2
) or Reduction of Nitrates
Nitrite Used to Cure Meats and Prevent Botulism
Can Be Reduced to Nitric Oxide in Hypoxic ConditionsIn Human Diet 80-90% from Reduction of Nitrates in VegetablesNitrates in Vegetables From Fertilizers or Plant StressesNitrite Readily Forms Cancer-Causing Nitrosamines in Stomach Acid Nitrites Oxidize Hemoglobin’s Iron From Ferrous (II) to Ferric(III) State - Unable to Carry Oxygen - Can be SeriousSlide19
Nitrosamines
Nitrosamines Produced by Reaction of Nitrites and Secondary Amines, Such as Proline
Strong Acids (Stomach) or High Temperatures of Frying Favor Production
Found in Processed Meats, Beer, Cigarette Smoke, Chewing Tobacco
Formation Inhibited by Vitamin C
Nitrosamine
NO
+
+ →
H
Secondary
Amine
Nitrite
H
2
NO
2
+
→ H
2
O + NO
+
Nitrosonium Ion
2H
+
Nitrous Acid IonSlide20
Nitrosamines
Nitrosamines Form DNA Adducts and Cause Cancer in Many Animal Species
Likely Carcinogens In Humans
Evidence for Gastric and Esophageal Cancer Risk
Nitrosamines in Tobacco Form From Nicotine
NNK is Nicotine Derived and Important in Carcinogenesis NNK in Tobacco and E-cigarettesNNK Activation by P-450 Activated Signaling Cascades & Uncontrolled Growth
Nicotine-derived nitrosamine ketone (NNK)
(4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone)Slide21
Reactive Nitrogen Species
Reactive Nitrogen Species Can Arise from any of the Molecules Described Here
The Most Potent Reactive Nitrogen Species is PeroxynitriteSlide22
Reactive Nitrogen Species
Peroxynitrite is Formed from Nitric Oxide and Superoxide
Peroxynitrite Can Readily React with DNA and Protein, Causing Damage
Cysteine Side Chains are Most Easily Oxidized
Tyrosine Side Chains of Proteins Can Be Nitrosylated
Transition Metals, Such as in Hemoglobin, Myoglobin, and Cytochromes Can Be Oxidized·NO + O
2·− ONOO
−
Nitric Oxide
Superoxide
PeroxynitriteSlide23
Amino Acid Metabolism
Introduction
There are 20 Common Amino Acids in Proteins Plus One Rare One
No One Single Pathway for Amino Acid Metabolism
Synthesis Pathways are Grouped According to Common Anabolic Precursors
α-ketoglutarateSerineAspartateAromatic
PyruvateHistidine
Essential Amino Acids Must Be in DietNon-Essential Ones Can be Made by OrganismEssential vs Non-Essential Varies in HumansSlide24
α-ketoglutarate Family
Transamination Plays an Important Role
α-keto Acid #1 +
Amino Acid X
Amino Acid #1
+ α-keto Acid XSlide25
α-ketoglutarate Family
Transamination to Make Glutamate
α-ketoacid #1
Amino Acid #2
α-keto Acid #1 +
Amino Acid X
Amino Acid #1
+ α-keto Acid X
Amino Acid X
α-ketoacid XSlide26
α-ketoglutarate Family
Glutamine Synthesis Uses Glutamine SynthetaseSlide27
α-ketoglutarate Family - Arginine Synthesis
Four Pathways to Make it
Deficiency of the Enzyme Arginase Leads to the Genetic Disease of Argininemia - Accumulation of Arginine and NH
4
+
in the Blood
ADMA
Arginine
Arginine
Citrulline + Aspartate
ATP
Argininosuccinate
AMP + 2 P
i
Ornithine + Urea
H
2
O
Citrulline + Nitric Oxide + H
2
O
NADP
+
NADPH + O
2
Demythlation
ADMASlide28
Serine Family
3-PG + NAD
+
3-phosphohydroxypyruvate
O-phosphoserine
Serine + P
i
H
2
O
Two Main Paths Lead to Serine
1. From 3-phosphoglycerate
(Connection to Glycolysis)
Glutamate
α-ketoglutarateSlide29
Serine Family
2. Exchanging Carbon with Glycine and Folates
(Important for Folate Recycling)
Serine + Tetrahydrofolate
Glycine + N
5
,N
10
-Methylene Tetrahydrofolate + H
2
OSlide30
Serine Family
Cysteine Metabolism
Multiple Ways of Making Cysteine
Primary Means Tied to Methionine Catabolism
Methionine
SAM
SAH
Homocysteine
Cystathionine
Cysteine
Serine
ATP
P
i
+ PP
i
Acceptor
CH
3
-Acceptor
Adenosine
H
2
O
β-ketobutyrate
Transmethylase
S-adenosylhomocysteine Hydrolase
Cystathionine β-synthase
Methionine Adenosyltransferase.
Cystathionase
Deficiency Leads to Homocystinuria
High Blood Levels - Cardiovascular Disease, Stroke Risk
H
2
O
NH
4
+Slide31
Serine Family
O-acetyl-L-serine
Acetyl-CoA
CoA-SH
L-cysteine + Acetate
H
2
S
2 Cysteine
2 NADH + 2H
+
2 NAD
+
Serine
L-cystine
L-cysteic Acid
L-cysteine
H
2
S
Sulfite
Other Cysteine MetabolismSlide32
Serine Family
Selenocysteine Metabolism
Sometimes Called 21st Amino Acid
Not Specified Directly in Genetic Code
Uses Stop Codon with Unusual Structure
Synthesized from Serine on tRNASerine + tRNA
SER
tRNA
Non-SER tRNA
SEL-A
SEL-D
SEL
tRNA
Incorporation Into ProteinsSlide33
Aspartate Family
All Family Members Arise from Aspartate
Aspartate Can be Made from One of Them - Asparagine
Numerous Paths Lead to Aspartate
Asparagine + H
2
O
Glutamate + Oxaloacetate
Argininosuccinate + AMP
α-ketoglutarate + Aspartate
Aspartate + NH
4
+
Aspartate + Citrullyl-AMP
Transamination
Hydrolysis
Reversal of
Reaction
Toxic
Urea CycleSlide34
Aspartate Family
Asparagine Metabolism
Aspartate + Glutamate + ATP
Asparagine + α-ketoglutarate + AMP + PP
i
Energetically Costly
Essentially Not Reversible
Asparagine Synthetase
Synthesis
Breakdown
Asparagine + H
2
O
Aspartate + NH
4
+
Toxic
AsparaginaseSlide35
Aromatic Family Outline
Tryptophan
Melatonin
Serotonin
Niacin
AuxinsPhenylalaninePhenylketonuriaTyrosineCatecholaminesThyroid Hormones
MelaninSlide36
Aromatic Family
Tryptophan, Phenylalanine, and Tyrosine
Each Derived from Phosphoenolpyruvate and Erythrose-4-phosphate
Synthesis Pathways Complex
Each Involves Shikimic Acid and Chorismic Acid
Phenylalanine and Tyrosine Pathways OverlapHormones and Neurotransmitters Made from EachSlide37
Aromatic Family
Tryptophan
Interesting Regulation of Synthesis in Bacteria
Attenuation - All 5 Genes on One Operon
When Tryptophan High, Transcription of Operon Aborts Early
When Tryptophan Low, Transcription of Operon Continues Through All GenesMolecules Made from TryptophanMelatoninCircadian Rhythm SensingAffects Mood, Sleep, Blood Pressure
Production Affected by Blue LightSerotonin
NeurotransmitterCauses Vasoconstriction
Enhances Memory/Learning, Contributor to Happy FeelingsNiacinVitamin B3Nicotinamide Derived From it - Part of NAD+/NADH & NADP+/NADPH
Deficiency Leads to Pellagra
AuxinsIndole-3-Acetic Acid Most ImportantStimulate Cell Division and Rooting in Plants
Melatonin
Serotonin
Indole-3-Acetic Acid
NiacinSlide38
Aromatic Family
Phenylalanine (PHE)
An Essential Amino Acid and Precursor of Tyrosine
PHE Hydroxylase Catalyzes Formation of Tyrosine from PHE
Deficiency of the Enzyme PHE Hydroxylase Causes Phenylketonuria
High PHE Levels Cause Damage to BrainTreatable by Reducing PHE LevelsNutrasweet Contains PHESlide39
Aromatic Family
Tyrosine (TYR)
Not Essential if PHE Present
Precursor of Catecholamines - L-Dopa, L-Dopamine, Norepinephrine, and Epinephrine
Donates Electrons to Reduce Chlorophyll in Photosystem II
Forms Radical in Ribonucleotide Reductase
CO
2
L-Norepinephrine
L-Norepinephrine
L-EphinephrineSlide40
Aromatic Family
Tyrosine Metabolism
L-Dopa
Precursor to Dopamine
Crosses Blood-Brain Barrier
Used to Treat Parkinson’s DiseaseDopamineNeurotransmitterInhibits Norepinephrine Release in Blood Vessels - Acts as VasodilatorReduces Insulin Production in PancreasDeficiency Causes Parkinson’s DiseaseLinks to Schizophrenia and ADHD
Norepinephrine
Hormone and NeurotransmitterWorks Through Noradrenergic ReceptorsFight or Flight Response
Increases Heart Rate and Blood PressureEpinephrine (Adrenalin)HormoneActions Similar to Norepinephrine
Fight or Flight ResponseIncreases Heart Rate and Blood PressureSlide41
Aromatic Family
Tyrosine is a Precursor of Thyroid Hormones
Secretion of Thyroglobulin
Export from Cell
Iodide Export
& Oxidation
Iodination
Transport Into Cell
Thyroglobulin Breakdown
Transport Into BloodSlide42
Aromatic Family
Tyrosine Metabolism
Thyroid Hormones
T3 (Triiodothyronine)
T4 (Thyroxine)
Deiodinases
All are Se-Containing Enzymes
More Active Form
More Abundant FormSlide43
Aromatic Family
Tyrosine Metabolism
Melanin - Oxidized and Polymerized Tyrosine
Benzoquinone Portion of Coenzyme Q
Tyrosine Unit
Further Polymerization
From TyrosineSlide44
Aromatic Family
Tyrosine Metabolism & Disease
Tyrosinemia - Problems with Tyrosine Catabolism
Type I
Type II
Type IIIAlcaptonuria - Black Urine DiseaseTreatmentsRestricted TYR/PHE DietLiver Transplant
Tyrosine
p-hydroxyphenylpyruvate
Homogentisate
4-Maleylacetoacetate
4-Fumarylacetoacetate
Fumarate + Acetoacetate
Tyrosine Transaminase
Type II
p-hydroxyphenylpyruvate Dioxygenase
Type III
Alcaptonuria
Alcaptonuria
4-fumarylacetoacetase
Type ISlide45
Pyruvate Family
Alanine Metabolism
Most Easily Produced from Pyruvate - Transamination
Byproduct of Catabolism of Valine, Leucine, and Isoleucine
Glucose-Alanine Cycle
Alanine Transaminase
Glutamate + Pyruvate
α-ketoglutarate + AlanineSlide46
Pyruvate Family
Leucine (LEU) /Valine (VAL) /Isoleucine (ILE) Metabolism
Branched Chain Amino Acids (BCAAs)
Several Common Steps
Start with Decarboxylation and Attachment of Two Carbon Piece to TPP
Valine and Leucine Pathways Involve Attachment of Two Carbon Piece to PyruvateIsoleucine Pathway Involves Attaching Two Carbon Piece to α-ketobutyratePenultimate Products - α-ketoisocaproate (LEU), α-ketoisovalerate (VAL), and α-keto-β-methylvalerate (ILE) Each is Transaminated to Make Final Amino Acid
Isoleucine
Leucine
ValineSlide47
Pyruvate Family
Leucine (LEU) /Valine (VAL) /Isoleucine (ILE) Metabolism
Synthesis Feedback Regulated Through Threonine Deaminase
Starting Material for ILE
Starting Material for VAL & LEU
Makes Starting
Material for ILE
Used by
All Three
High ILE
Favors VAL & LEU
High VAL
Favors ILESlide48
Pyruvate Family
Histidine (HIS) Metabolism
Most Complex of All the Amino Acids
Overlaps Nucleotide Metabolism with Ribose-5-Phosphate & PRPP
10 Steps in Pathway
Second Enzyme of Pathway (ATP-phosphoribosyltransferase) Feedback Inhibited by Histidine
HistidineSlide49
Amino Acid Catabolism
Glycogenic
Alanine, Cysteine, Glycine, Serine
Asparagine, Aspartate, Arginine
Histidine, Proline, Glutamine
Glutamate, Methionine, ValineKetogenicLysine, LeucineBoth
Threonine,Tryptophan, Tyrosine,
Phenylalanine, Isoleucine
Three Categories1. Glycogenic - Broken Down to Glycolysis/Gluconeogenesis Intermediates
2.
Ketogenic - Broken Down to Acetyl-CoA3. Both - Makes Intermediates in Both PathwaysSlide50
Amino Acid Catabolism
Most Diseases of Amino Acid Metabolism Arise from Problems with Catabolism
Alcaptonuria - Phenylalanine and Tyrosine
Methylmalonic Acidemia - Methionine
, T
hreonine, Isoleucine and ValineMaple Syrup Urine Disease - Valine, Leucine, IsoleucineHomocystinuria - MethionineTyrosinemia - TyrosineArgininemia - ArginineHypermethioninemia - MethionineHyperlysinemia - Lysine
Glycine Encephalopathy - Glycine
Propionic Acidemia - Methionine, Threonine, Isoleucine and
ValineHyperprolinemia - Proline