metabolisme asam amino

Chapter 20: Amino acid metabolism Takusagawas Note

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Chapter 20: Amino Acid Metabolism Amino acids from proteins are: - precursors of compounds - energy source (i.e., converted to acetyl-CoA, etc.) Amino acids are obtained in diet and/or turnover of cellular proteins. Major problem in amino acid degradation is elimination of amino group (-NH2) since NH3 from -NH2 is very toxic. Ammonia eliminations are: - Conversion to urea (mammals) - Conversion to uric acid (birds) Carbon skeleton of amino acid metabolism is: - NH2 group is removed by transamination & oxidative deamination to urea. Transamination

R1 C

O

COO- + R2 CH NH2

COO-

CH NH2

COO-

R1

C COO-O

R2+E-PLP

-KA2AA1 -KA1 AA2

CH2

O

C-O CH2 C CO

O-O

C

O

CH2-O CH2 CH C

OO-

H3N+

transaminase

Glutamate

-Ketoglutarate

-Keto acid

L-amino acid

R C

O

C

OO-

R CH

H3N+

C

OO- C

O-O CH2 CH C

OO-

H3N+

O

C-O CH2 C CO

O-O

transaminase

Aspartate

Oxaloacetate Then Asp Urea Oxidative deamination

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CH2

O

C-O CH2 C CO

O-O

C

O

CH2-O CH2 CH C

OO-

H3N+

NH3 glutamatedehydrogenase

NAD+ + H2O

NADH + H+

transaminase

glutamate

-ketoglutarate

-keto acid

L-amino acid

R C

O

C

OO-

R CH

H3N+

C

OO-

Urea


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Pyridoxal-5-phosphate (PLP) is co-enzyme (co-factor) of transaminase.

Aminotransferase reactions occur in two stages (Ping-Pong Bi Bi reaction): 1. Amino acid + Enzyme -Keto acid + Enzyme-NH2 2. -Ketoglutarate + Enzyme-NH2 Enzyme + Glutamate Details of aminotransferase reactions are shown in Fig. 24-2. Stage-0: Enzyme-PLP Schiff base formation PLP is covalently attached to the enzyme via a Schiff base linkage between aldehyde group

of PLP and Lys (-amino group) of enzyme. E-Lys + PLP E-PLP

N

C

O-

CH3H

H2C

H N

(CH2)4

H

Enzyme

2-O3PO

+

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Stage I: Conversion of an amino acid to -keto Acid 1. Transimination: Amino acids nucleophilic amino group attacks the E-PLP Schiff base

carbon atom in a transimination reaction to form E-PLP-AA. Then E-PLP-AA is E-Lys + PLP-AA.

E-PLP + AA [E-PLP-AA] E-Lys + PLP-AA

2. Tautomerization: AA-PLP tautomerizes to an -keto acid-PMP by the active-site Lys

catalyzed removal of the amino acid -hydrogen and protonation of PLP atom C4. AA-PLP -Keto acid-PMP

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3. Hydrolysis: -Keto acid-PMP is hydrolyzed to PMP and -Keto acid. -Keto acid-PMP + H2O -Keto acid + PMP

Stage II: Conversion of an -keto acid to an amino acid (reverse reactions of stage I) 3. -Keto acid + PMP -Keto acid-PMP 2. -Keto acid-PMP AA-PLP 1. E-Lys + PLP-AA [E-PLP-AA] E-PLP + AA Note: All amino acids form the E-PLP-AA intermediate: AA + E-PLP E-PLP-AA.

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NH2 E

N CH3H

O-O3P

HC

HNC COO

-H

CC

OH

E-PLP-AA is then converted by: 1. Transamination 2. Decarboxylation 3. Elimination from - or -carbon 4. Racemization (D L) 5. Others

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Urea Cycle Urea is formed from ammonia (NH3), amino group (NH2) of Asp, and bicarbonate (HCO3-) by

urea cycle in liver.

C

O

H2N NH2

NH3 NH2 of Asp

HCO3-

- Five enzymes are involved in urea synthesis in urea cycle. Two enzymes are in mitochondrion. Three enzymes are in cytosol. - Therefore, the urea cycle occurs partially in the mitochondrion and partially in the cytosol.

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Reactions in urea cycle 1. Carbamoyl phosphate synthetase (Regulating enzyme) Formation of carbamoyl phosphate from NH3 and HCO3- (bicarbonate) using ATP as energy

source. HCO3- + NH3 + 2ATP H2N-CO(OPO32-) + 2ADP + Pi

1st ATP

2nd ATP

2. Ornithine transcarbamoylase Transfer carbamoyl group (O=C-NH2) to ornithine to produce citrulline. Ornithine + O=C-NH2(PO32-) Citrulline + Pi 3. Argininosuccinate synthetase Acquisition of the second urea nitrogen atom from Asp. Citrulline + Asp Argininosuccinate

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4. Argininosuccinase Elimination of arginine from the aspartate carbon skeleton to form fumarate. Argininosuccinate Fumarate + Arginine 5. Arginase Hydrolysis of arginine to yield urea and regenerate ornithine. Arginine Urea + Ornithine Overall reaction of urea cycle is:

CO2 + NH3+ + 3ATP + Asp + 2H2O Urea + 2ADP + 2Pi + AMP + PPi ( 2Pi) + Fumarate

The urea cycle converts two amino groups (one from NH3 and one from Asp) and a carbon atom

(HCO3-) to non-toxic excretion product, urea, at the cost of 4 high-energy phosphate bonds (i.e., 4ATP).

However, oxidations of urea cycles substrate (Glu) and product (malate) produce 2 NADH (= 6

ATP) as shown in Fig. 24-7.

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The urea cycle is conjunct with apartate-argininosuccinate shunt of tricarboxylic acid (TCA) cycle as shown below. This is called Krebs bicycle. Note: tricarboxylic acid cycle = citric acid cycle = Krebs cycle.

Oxaloacetate is one of the most important precursor of:

Protein

UreaAsp

Gluconeogenesis

CAC (condenses with acetyl-CoA)

Oxaloacetate

Regulation of the urea cycle - is regulated by carbamoyl phosphate synthetase. - Carbamoyl phosphate synthetase is allosterically activated by N-acetylglutamate. Thus, N-

acetyl-glutamate plays an important role in urea cycle regulation. COO-

(CH2)2CH

-OOC

NH

C

O

CH3

8N-Acetyl-glutamate


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- Acetyl-Glu is synthesized by acetyl-glutamate synthase Glu + Acetyl-CoA N-acetyl-Glu. - N-acetyl-Glu formation can be as follows: 1. Breakdown of protein produces amino acids including Glu (i.e., [Glu] ). 2. Need urea cycle to be activated since amino acid degradation produces amines. 3. In the mean time, [Glu] causes [N-acetyl-Glu] 4. [N-acetyl-Glu] increases the activity of carbamoyl phosphate synthetase. Thus, urea

cycle is activated. Ammonia transport mechanism - Ammonia (NH3) is produced in all tissue, but the urea cycle is only carried out in liver.

Thus, NH3 must be transported to liver with non-toxic form. NH3 is converted to glutamine (Gln) which is not toxic. ATP + NH4+ + Glu ADP + Pi + Gln + H+

Glutamine synthetase

- Gln is hydrolyzed to Glu and NH4 in liver. Gln + H2O Glu + NH4+

glutaminase- NH4+ is converted to urea. Another special system between muscle and liver to get nitrogen to the liver: Glucose-alanine

cycle is shown below. - Amino group in Glu produced from amino acids NH3 in muscle is transferred to pyruvate. - The aminated pyruvate, Ala, is transported to liver where the NH2 is transported to -

ketoglutarate. - The aminated -ketoglutarate, Glu, releases NH3. - NH3 enters the urea cycle and is converted to urea. - In this glucose-alanine cycle, muscle uses glucose and excretes nitrogen, whereas liver

converts alanine to glucose and excretes NH3 in urea cycle.

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Amino acids skeleton metabolism

Keto Keto & Gluco

Gluco

Leu Lys

Ile Thr Phe Try Trp

Ala Cys Gly Ser Asp Asn Met Val Arg Glu Gln His Pro

- 20 amino acids are converted to 7 common intermediates. Those are:

Both glucogenic and ketogenic intermediate (do not confuse!)

Ketogenic intermediates (form ketone bodies)

Glucogenic intermediates (form glucose)

1. Pyruvate

2. -ketoglutarate3. Succinyl-CoA4. Fumarate5. Oxaloacetate

6. Acetyl-CoA7. Acetoacetate

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Example of Amino acid degradation Alanine, Cysteine, Glycine, Serine, and Threonine are degraded to Pyruvate - Degradations of these amino acids involve: 1. Elimination of -NH2, -OH, -SH 2. Transfer of hydroxymethyl group 3. Oxidation-reduction - Pathways are shown Fig. 24-9.

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Amino acid biosynthesis Tetrahydrofolate Cofactors: Metabolism of C1 Units - Tetrahydrofolate (THF) functions to transfer C1 units in several oxidation states. - Most reactions require NADPH/NADH. - THF is composed of three units: 2-Amino-4-oxo-6-methylpterin p-Aminobenzoic acid Glutamates

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- THF is derived from folic acid (one of vitamin) by two-stage reduction. Both reactions are catalyzed by dihydrofolate reductase (DHFR).

- Inhibition of DHFR inhibits nucleic acid synthesis since THF transfers C1 units to biosyntheses of proteins and nucleic acids.

- N5 and N10 in THF are important nitrogens, since C1 units are covalently attached to THF at

its positions 5N, 10N, or both 5N and 10N. - C1 units are listed in Table 1.

- The C1 units carried by THF are interconverted to: Methionine Thymidylate (dTMP) Formylmethionine-tRNA Purines

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Reactions involved in THF are oxidation-reduction, cyclization and hydrolysis

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Sulfonamides competitively inhibit bacterial synthesis of THF

H2N S

O

O

NH R H2N C

O

OH

Sulfonamides(R = H sulfanilamide)

p-Aminobenzoic acid

Why? Because sulfonamides are: - structural analogs of p-aminobenzoic acid constituent of THF. - antibiotics (sulfa drugs) which competitively inhibit bacterial synthesis of THF. Amino acid biosynthesis and related products Amino acids are not only the components of proteins, but also precursors to various compounds

including neurotransmitters, hormones and porphyrins. Essential and nonessential amino acids in humans

Essential amino acids --- Amino acids that are not synthesized in human bodies. - Plants and microorganisms can make essential amino acids.

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Nonessential amino acids --- Amino acids that are synthesized in human bodies. - These amino acids are synthesized from intermediates of glycolysis and the citric acid cycle.

Asparagine

Aspartate

Proline

Glutamine

Glutamate

Alanine

Cystine

GlycineSerine

C.A.C.

Oxaloacetate

-Ketoglutarate

Pyruvate

Glucose-6-phosphate

Fructose-6-phosphate

Triose-3-phosphate

Glycerate-3-phosphate

Phosphoenolpyruvate

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Details of syntheses of Ala, Asp, Glu, Asn, and Gln are shown in Fig. 24-41.

Donor amino group

Glutamine synthetase is a central control point in nitrogen metabolism, since glutamine is the

amino group donor in the formation of many biosynthetic products as well as being a storage form of ammonia.

- is 12 subunits protein (bacteria). - is inhibited by two mechanisms: 1. Feedback inhibition. (In general, the final product inhibits the first reaction) - His, Try, carbamoyl phosphate, AMP, CTP, glucosamine-6-phosphate which are all end

products of pathways leading from glutamine (i.e., receive amide nitrogen from glutamine) are allosteric inhibitors.

- Ala, Ser, Gly inhibit by reflecting the cells high nitrogen level, i.e., Ala, Ser and Gly are synthesized only the citric acid cycle is saturated.

- When the citric acid cycle is saturated, biosyntheses are started.

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2. Covalent modification. - Adenylylation - deadenylylation and uridylylation - deuridylylation Under conditions of nitrogen excess: 1. High [glutamine] activates uridylyl-removing enzyme. 2. Uridylyl-removing enzyme catalyzes deuridylylation of adenylyltransferase (PII-4UMP

PII). 3. Under a high [glutamine/-ketoglutarate] ratio, the PII catalyzes adenylylation of glutamine

synthetase, and inactivates it. Under conditions of nitrogen limitation: 1. High [-Ketoglutarate] activates uridylyltransferase. 2. Uridylyltransferase catalyzes uridylylation of adenylyltransferase (PII PII-4UMP). 3. The uridylylated adenylyltransferase (PII-4UMP) catalyzes deadenylylation of glutamine

synthetase, and activates it. 4. Activated glutamine synthetase catalyzes glutamate to glutamine reaction.

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A specific tyrosine residue of adenylyltransferase (PII) is: - uridylylated by uridylyltransferase (PII-4UMP is an active for deadenylylation). - deuridylylated by uridylyl-removing enzyme (PII is an active for adenylylation). Similarly, a specific tyrosine residue of glutamine synthetase is: - adenylylated by deuridylylated adenylyltransferase (PII). Adenylylated enzyme is inactive. - deadenylated by uridylylated adenylyltransferase (PII-4UMP). Deadenylylated enzyme is

active.

Glutamine

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Glutamate is the precursor of proline, ornithine, and arginine

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Serine, cysteine, and glycine are derived from 3-phosphoglycerate

Gly Cys

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S-adenosylmethionine (SAM) is synthesized from methionine and ATP - SAM is the major methyl group donor molecule, and by releasing the methyl group, SAM

becomes S-adenosylhomocysteine.

Donor methyl group

- High level of homocysteine is one of the risk factors for coronary heart disease (heart

attack).

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Glycine is synthesized from serine by removing CH2OH group.

5,10-methylene THFTHF

Serine hydroxymethyl transferase

Glycine L-Serine

Tyrosine is synthesized from phenylalanine

NADP+ NADPH + H+

dihydrobiopterinO2 + tetrahydrobiopterin

Phenylalanine-4-monooxygenase

L-TyrL-Phe

- Genetic disease, phenylketonuria is caused by less active or inactive phenylalanine-4-

monooxygenase. This disease produces abnormal level of phenylpyruvate in urine, since phenylalanine is converted to phenylpyruvate instead of L-tyrosine.

Phe Tyr

Phenylpyruvate Amino acids are precursors of porphyrins, amines and peptides (glutathione) Porphyrin synthesis Porphyrins are derived from succinyl-CoA and glycine Gly + Succinyl-CoA -Aminolevulinate (ALA) + CO2 + CoASH - PLP is involved in the catalytic reaction. - Pyrrole ring is the product of two ALA molecules. 2ALA Porphobilinogen (PBG) Uroporphyrinogen III is synthesized from four PBGs. 4PBG Hydroxymethylbilane Uroporphyrinogen III

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Overall heme biosynthesis is taken place in both mitochondrion and cytosol.

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There are several genetic defects in heme biosynthesis: 1. Uroporphyrinogen III cosynthase deficiency = congenital erythropoietic porphyria Red urine, reddish teeth, photosensitive skin, increased hair growth. 2. Ferrochelatase deficiency = erythropoietic porphyria Amine synthesis (Mainly decarboxylation by PLP dependent enzymes) Some of amines are important neurotransmitters and hormones. Biosynthesis of -aminobutyric acid (GABA, neurotransmitter), histamine (allergic response),

and serotonin (neurotransmitter)are shown below.

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Epinephrine, norepinephrine and dopamine biosyntheses

HO

HO C

X

H

CH2 NH R

X = OH, R = CH3 EpinephrineX = OH, R = H NorepinephrineX = H, R = H Dopamine

- Tyrosine is the precursor of these hormones.

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Parkinsons disease


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- Deficiency in dopamine production is associated with Parkinsons disease (deficiency of tyrosine hydroxylase). L-DOPA has been used to treat Parkinsons disease. - In melanocytes:

Tyr tyrosinase

O2 H2O

DOPA

H2OO2

tyrosinase

O

O

CH2 CH

COO-

NH3+

phenyl-3,4-quinone

Melanine (black skin pigment)polymerization

- Tyrosine hydroxylase (tyrosinase) is an important enzyme. Glutathione

- Important functions of GSH is elimination H2O2 and reduction of protein thiol-disulfied.

ProteinS

S ProteinSH

SH2GSH GSSG

thiol transferase

Amino acids skeleton metabolism

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metabolisme asam amino