Amino acid metabolism I Jana Novotná Department of the Medical Chemistry and Clinical Biochemistry The 2nd Faculty of Medicine, Charles Univ.

Amino acid metabolism I

Jana NovotnáDepartment of the Medical Chemistry and Clinical Biochemistry

The 2nd Faculty of Medicine, Charles Univ.

Metabolic relationship of amino acids

Body proteins

Proteosynthesis Degradation

Amino acidpoolDietary

proteins

Digestion

Transaminatio

n

NONPROTEINDERIVATIVESPorphyrinsPurinesPyrimidinesNeurotransmittersHormonesKomplex lipidsAminosugars

UREA NH3

Carbon skeletonconversion

250 – 300 g/day

acetyl CoASaccharideslipides

CO2

H2O

GlycolysisKrebs cycle

Conversion to

Ketonbodies

Digestive tractEndopeptidases – hydrolyse the peptide bond inside a chain: pepsin (stomach), trypsin, chymotrypsin, elastase (pancreas)

Exopeptidases – split the peptide bond at the end of a protein molecule:

aminopeptidase, carboxypeptidases, dipeptidases (small intestine)Hydrolysis of proteins polypeptides oligopeptides amino acids intestinal lumen transport to target tissuesPepsin (pH 1.5 – 2.5) – hydrolysis of peptide bond before Tyr, Phe and between Leu and Glu.Trypsin (pH 7.5 – 8.5) – peptide bond after Lys a Arg.Chymotrypsin (pH 7.5 – 8.5) – peptide bond after Trp, Phe,Tyr, Met, Leu.Pancreatic elastase (pH 7.5 – 8.5) - peptide bond after Ala, Gly and Ser

Degradation of amino acids intracellularly the first step is deamination, transamination, oxidative decarboxylation

Enzymes cleaving the peptide bond

Absorption of amino acids

• Absorption from the lumen of small intestine by transepitelial transport

• Semispecific Na+-dependent transport system

• Na+-dependent carriers transport both Na+ and an amino acid.

• At least six different Na+-dependent carriers:

- neutral AA- proline and hydroxyproline- acidic AA- basic AA (Lys, Arg) and cistine

Clinical note:

Genetically determined defect in the transport of amino acids across the brush border membranes of cells in both small intestine and renal tubules

Cystinuria – AR disease, caused by mutation in two genes for transporter proteins in the kidney proper reabsorption of basic, or positively charged, amino acids (Lys, Arg and ornithine) and cysteine into bloodstream is prevent Cys is oxidized to insoluble cystine formation of kidney stones renal colic.

Hartnup disease – relatively rare AR disease – defect in tranport of neutral AA including essential (Ile, Leu, Val, Phe, Thr, Trp - availability of essential AA may cause a variety clinical disorders

The urine of newborns is routinely screening.

g-Glutamyl cycle and amino acid transport

Gamma-glutamyl transferase (gamma-glutamyl transpeptidase, GGT)

Found in many tissues, mainly in the liver.

Diagnostic marker for liver disease - elevations in GGT in patients with chronic viral hepatitis infections.

Transport of AA across cell membrane by reacting with glutathion to for g-glutamyl amino acid

AA is released into the cell. Glutathion is resinthesized.

Transamination - exchange of NH2 group with C=O

General reactions of amino acid catabolism

General reactions of amino acid catabolism

Deamination

General reactions of amino acid catabolism

Decarboxylation

Decyrboxylation of AA gives amines having a variety of functions.

Transamination reaction

The first step in the catabolism of most amino acids is removal of a-amino groups by enzymes transaminases

or aminotransferases

All aminotransferases have the same prostethic group and the same reaction mechanism.

The prostethic group is pyridoxal phosphate (PPL), the coenzyme form of pyridoxine (vitamin B6)

Active metabolic form of vitamin B6

Mechanism of transamination reaction: PLP complex with enzyme accept an amino group to form pyridoxamine phosphate, which can donate its amino group to an a-keto acid.

All amino acids except threonine, lysine, and proline can be transaminated

Transaminases are differ in their specificity for individual L-amino acid. The enzymes are named for the amino group donor.

Clinically important transaminases

ALT

Alanine transaminase ALT(previously called serum glutamate-pyruvate transaminase – SGPT)Predominantly found in the liver. Important in the diagnosis of liver (viral hepatitis drug toxicity), ALT is a more specific indicator of liver inflammation than AST.

Aspartate transaminase AST(previously called serum glutamate-oxaloacetate transaminase – SGOT).- Found in the liver, heart, skeletal muscles, kidneys, brain, and red blood cells. - Elevated in liver diseases, myocardial infarction, acute pancreatitis,

acute hemolytic anemia, severe burns, acute renal diseases, musculoskeletal diseases, and trauma (in 1954 defined as a biochemical marker for the diagnosis of acute myocardial infarction)

Deamination

Amino acids FMN H2O+ +

a-keto acids FMNH2 NH3

L-amino acid oxidase

A. Oxidative deamination

FMN

H2O2 H2O + O2

+ +O2

catalse

B. Nonoxidative deamination

serine

pyruvate

threonine

a-ketobutyrate

+ +

Serin-threonin dehydratase

• L-amino acid oxidase produces

ammonia and a-keto acid directly,

using FMN as cofactor.• The reduced form of flavin must

be regenerated by O2 molecule.

• This reaction produces H2O2

molecule which is decompensated

by catalase.

Is possible only for hydroxy amino acids

NH3 + H2O NH3 + H2O

Decarboxylation

• process is catalysed by enzymes decarboxylase – cofactor is pyridoxalphosphate

• R-CHNH2-COOH R-CH2NH2 + CO2

• takes place only in small quantities • primary amines

• biologically active amines• hormones (neurotransmitters, coenzymes)

Biogenic amines

• Tyrosine norepinephrine, epinephrine, dopamine, DOPA

• Histidine histamine• Tryptophane serotonine, melatonine• Ornithine putrescine, polyamines• Glutamate GABA• Serine ethanolamine

Synthesis of non-essential amino acids

Overview of the synthesis of non-essential amino acids

The carbon of 10 AA may be produced from glucose through intermediates of glycolysis or the TCA cycle.Tyrosine from phenylalanine.The sulphur of cysteine – from methionine.

Amino acids derived from intermediates of glycolysis

The major pathways for serine synthesis from glucose and serine degradation

Glycine biosynthesis from serine

Reaction involves the transfer of the hydroxymethyl group from serine to the cofactor tetrahydrofolate (THF), producing glycine and N5,N10-methylene-THF.

Copy from: http://themedicalbiochemistrypage.org/amino-acid-metabolism.html

Glycine oxidation to CO2

Glycine produced from serine or from the diet can also be oxidized by glycine decarboxylase (also referred to as the glycine cleavage complex, GCC) to yield a second equivalent of N5,N10-methylene-tetrahydrofolate as well as ammonia and CO2.

Copy from: http://themedicalbiochemistrypage.org/amino-acid-metabolism.html

Tetrahydrofolate acts as a carrier of reactive single C units

Copy from: http://www.chembio.uoguelph.ca/educmat/chm452/lectur25.htm

Serine glycine – formation of N5,N10-methylen THFGlycine CO2 - formation of N5,N10-methylen THFHomocysteine methionine – donor of C is N5-methyl THFHistidine degradation – formation of N5-formiminoTHF; N5,N10-metnhenyl a N10-formyl THFTryprophane degradation – formation of N10-formyl THF

Metabolism of glycine

Cysteine synthesis

Copy from: http://themedicalbiochemistrypage.org/amino-acid-metabolism.html

1. Conversion of SAM to homocysteine.

2. Condensation of homocysteine with serine to cystathione.

3. Cystathione is cleavaged to cysteine.

Conversion of homocysteine back to Met. N5-methyl-THF is donor of methyl group.

*

*folate + vit B12

Homocystinuria Genetic defects for both the synthase and the lyase.

Missing or impaired cystathionine synthase leads to homocystinuria.High concentration of homocysteine and methionine in the urine.

Homocysteine is highly reactive molecule.

Disease is often associated with mental retardation, multisystemic disorder of connective tissue, muscle, CNS, and cardiovascular system.

Clinical note

Relationship between glutamate, glutamine and a-ketoglutarate

a-ketoglutarate glutamate glutamine

NH3

NH3

NH3

NH3

Glutamate + NAD+ + H2O a-ketoglutarate NH3+ + NADH

Glutamate NH3+ glutamine

ATP ADP

Glutamine H2O+ glutamate NH3+

A. Glutamate dehydrogenase

B. Glutamine synthetase (liver)

C. Glutaminase (kidney)

From transamination reactions

To urea cycle

Amino acid degradation

Degradation of AA

20 amino acids are converted to 7 products:

pyruvate acetyl-CoA acetoacetate a-ketoglutarate succinyl-CoA oxalacetate fumarate

Glucogenic amino acids

formed: a-ketoglutarate, pyruvate, oxaloacetate, fumarate, or succinyl-CoA

Aspartate Asparagine Arginine Phenylalanine Tyrosine Isoleucine

Methionine Valine Glutamine Glutamate Proline Histidine

Alanine Serine Cysteine Glycine Threonine Tryptophan

Ketogenic amino acids

formed acetyl CoA or acetoacetate

Lysine

Leucine

Both glucogenic and ketogenic amino acids

formed: a-ketoglutarate, pyruvate, oxaloacetate, fumarate, or succinyl-CoA in

addition to acetyl CoA or acetoacetate

IsoleucineThreonineTryptophanPhenylalanineTyrosine

Amino acids that form acetyl-CoA and acetoacetate

Amino acids related through glutamate

Synthesis and degradation of proline

Histidine degradation

Amino acids that form succinyl-CoA

Amino acids related to oxalacetate

Aspartate and asparagine

The sulfur for cysteine synthesis comes from the essential amino acid methionine.

SAM serves as a precurosor for numerous methyl transfer reactions (e.g. the conversion of norepinephrine to epinenephrine).

Cysteine and methionine are metabolically related

Condensation of ATP and methionine yield S-adenosylmethionine (SAM)

SAM

valine isoleucine leucine

a-ketoglutarate glutamate (transamination)

a-ketoisovalerate a-keto-b-methylbutyrate a-ketoisokaproate

oxidative decarboxylationDehydrogenase of a-keto acids*

CO2

NAD+

NADH + H+

isobutyryl CoA a-methylbutyryl CoA isovaleryl CoA

Dehydrogenation etc., similar to fatty acid b-oxidation

propionyl CoA acetyl CoA

acetoacetate

acetyl CoA

propionyl CoA+ +

Degradation of branched amino acids

Branched-chain aminoaciduriaDisease also called Maple Syrup Urine Disease (MSUD) (because of the characteristic odor of the urine in affected individuals).

Deficiency in an enzyme, branched-chain α-keto acid dehydrogenase leads to an accumulation of three branched-chain amino acids and their corresponding branched-chain α-keto acids which are excreted in the urine.

There is only one dehydrogenase enzyme for all three amino acids.

Mental retardation in these cases is extensive.

Clinical note

Biosynthesis of tyrosine from phenylalanine

Phenylalanine hydroxylase is a mixed-function oxygenase: one atom of oxygen is incorporated into water and the other into the hydroxyl of tyrosine. The reductant is the tetrahydrofolate-related cofactor tetrahydrobiopterin, which is maintained in the reduced state by the NADH-dependent enzyme dihydropteridine reductase

Tetrahydrobiopterin as a cofactor of hydroxylases

Dihydrobiopterin

• Hyperphenylalaninemia, phenylketonuria - complete deficiency of phenylalanine hydroxylase (plasma level of Phe raises from normal 0.5 to 2 mg/dL to more than 20 mg/dL).

• The mental retardation is caused by the accumulation of phenylalanine, which becomes a major donor of amino groups in aminotransferase activity and depletes neural tissue of α-ketoglutarate.

• Absence of α-ketoglutarate in the brain shuts down the TCA cycle and the associated production of aerobic energy, which is essential to normal brain development.

• Newborns are routinelly tested for blood concentration of Phe.

• The diet with low-phenylalanine diet.

Clinical note

Tryptophan catabolism

Tryptophan has complexcatabolic pathway: 1. the indol ring is

ketogenic2. the side chain alanin

gluconeogenesis

Xanthurenic acid isexcrete in the urine.Nicotinamide NAD andNADP.

Enzymes which metabolised amino acides containe vitamines as cofactors

THIAMINE B1 (thiamine diphosphate) oxidative decarboxylation of a-ketoacids

RIBOFLAVIN B2 (flavin mononucleotide FMN, flavin adenine dinucleotide FAD)oxidses of a-amino acids

NIACIN B3 – nicotinic acid (nikotinamide adenine dinucleotide NAD+

nikotinamide adenine dinukleotide phosphate NADP+)dehydrogenases, reductase

PYRIDOXIN B6 (pyridoxalphosphate)transamination reaction and decarboxylation

FOLIC ACID (tetrahydropholate)Meny enzymes of amino acid metabolism

Pictures were taken from textbooks:

Marks´ Basic Medical Biochemistry A Clinical Approach. Four edition M. Lieberman, A.D. Marks ed., 2013. Essentials of Medical Biochemistry With Clinical Cases. First edition. N.V. Bhagavan, Chung-Eun Ha ed., 2011.