Amino acid metabolism I

Transcription

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

2 Metabolic relationship of amino acids Body proteins Dietary proteins Glycolysis Krebs cycle Proteosynthesis g/day Amino acid pool Degradation Carbon skeleton conversion UREA NH 3 Conversion to NONPROTEIN DERIVATIVES Porphyrins Purines Pyrimidines Neurotransmitters Hormones Komplex lipids Aminosugars Carbohydrates Lipids Acetyl CoA CO 2 H 2 O Ketonbodies

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4 Enzymes cleaving the peptide bond Digestive tract: Endopeptidases hydrolysis of peptide bond inside a polypeptide 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 tissues Pepsin (ph ) hydrolysis of peptide bond before Tyr, Phe and between Leu and Glu. Trypsin (ph ) peptide bond after Lys a Arg. Chymotrypsin (ph ) peptide bond after Trp, Phe,Tyr, Met, Leu. Pancreatic elastase (ph ) - peptide bond after Ala, Gly and Ser Degradation of amino acids intracellularly the first step is deamination, transamination, oxidative decarboxylation

5 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

6 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.

7 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.

8 General reactions of amino acid catabolism Transamination - exchange of NH 2 group with C=O

9 General reactions of amino acid catabolism Deamination

10 General reactions of amino acid catabolism Decarboxylation Decarboxylation of AA gives amines having a variety of functions.

11 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 B 6 )

12 Active metabolic form of vitamin B6

13 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.

14 All amino acids except threonine, lysine, and proline can be transaminated. Transaminases are differ in their specificity for individual L-a-amino acid. The enzymes are named for the amino group donor.

15 Clinicaly important transaminases 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) ALT

16 A. Oxidative deamination Amino acids + FMN + H 2 O a-keto acids + FMNH 2 + NH 3 Deamination L-a-amino acid oxidase FMN O 2 catalase. catalse H 2 O 2 H 2 O + O 2 L-a-amino acid oxidase produces ammonia and a-keto acid directly, using FMN as cofactor. The reduced form of flavin must be regenerated by O 2 molecule. This reaction produces H 2 O 2 molecule which is decompensated by B. Nonoxidative deamination serine Reaction is possible only for hydroxy amino acids threonine Serin-threonin dehydratase pyruvate a-ketobutyrate + + NH 3 + H 2 O NH 3 + H 2 O

17 Decarboxylation process is catalysed by enzymes decarboxylase cofaktor is pyridoxalphosphate R-CHNH 2 -COOH R-CH 2 NH 2 + CO 2 takes place only in small quantities primary amines biologically active amines hormones (neurotransmitters, coenzymes)

18 Synthesis of non-essential amino acids

19 Overview of the synthesis of nonessential 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.

20 Amino acids derived from intermediates of glycolysis

21 The major pathways for serine synthesis from glucose and serine degradation

22 Glycine biosynthesis from serine Reaction involves the transfer of the hydroxymethyl group from serine to the cofactor tetrahydrofolate (THF), producing glycine and N 5,N 10 -methylene-thf. Copy from:

23 Glycine oxidation to CO 2 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 N 5,N 10 -methylene-tetrahydrofolate as well as ammonia and CO 2. Copy from:

24 Tetrahydrofolate acts as a carrier of reactive single C units Serine glycine formation of N 5,N 10 -methylen THF Glycine CO 2 - formation of N 5,N 10 -methylen THF Homocysteine methionine donor of C is N 5 -methyl THF Histidine degradation formation of N 5 -formiminothf; N 5,N 10 -metnhenyl a N 10 -formyl THF Tryprophane degradation formation of N 10 -formyl THF Copy from:

25 Metabolism of glycine

26 Cysteine synthesis Conversion of homocysteine back to Met. N 5 - methyl-thf is donor of methyl group. * *folate + vit B Conversion of SAM to homocysteine. 2. Condensation of homocysteine with serine to cystathione. 3. Cystathione is cleavaged to cysteine. Copy from:

27 Clinical note 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.

28 Relationship between glutamate, glutamine and a-ketoglutarate NH 3 NH 3 a-ketoglutarate glutamate glutamine NH 3 NH 3 A. Glutamate dehydrogenase Glutamate + NAD + + H 2O a-ketoglutarate + NH 3 + NADH From transamination reactions To urea cycle B. Glutamine synthetase (liver) ATP ADP Glutamate + NH 3 glutamine C. Glutaminase (kidney) Glutamine + H 2 O glutamate + NH 3

29 Amino acid degradation

30 Degradation of AA 20 amino acids are converted to 7 products: pyruvate acetyl-coa acetoacetate a-ketoglutarate succinyl-coa oxalacetate fumarate

31 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

32 Ketogenic amino acids formed acetyl CoA or acetoacetate Lysine Leucine

33 Both glucogenic and ketogenic amino acids formed: a-ketoglutarate, pyruvate, oxaloacetate, fumarate, or succinyl-coa in addition to acetyl CoA or acetoacetate Isoleucine Threonine Tryptophan Phenylalanine Tyrosine

34 Amino acids that form acetyl-coa and acetoacetate

35 Amino acids related through glutamate

36 Synthesis and degradation of proline

37 Histidine degradation

38 Amino acids that form succinyl-coa

39 Amino acids related to oxalacetate Aspartate and asparagine

40 Cysteine and methionine are metabolically related The sulfur for cysteine synthesis comes from the essential amino acid methionine. SAM Condensation of ATP and methionine yield S-adenosylmethionine (SAM) SAM serves as a precurosor for numerous methyl transfer reactions (e.g. the conversion of norepinephrine to epinenephrine).

41 Degradation of branched amino acids valine isoleucine leucine a-ketoglutarate glutamate (transamination) a-ketoisovalerate a-keto-b-methylbutyrate a-ketoisokaproate CO 2 oxidative decarboxylation Dehydrogenase of a-keto acids* NAD+ NADH + H + isobutyryl CoA a-methylbutyryl CoA isovaleryl CoA Dehydrogenation etc., similar to fatty acid b-oxidation propionyl CoA acetyl CoA acetyl CoA + + propionyl CoA acetoacetate

42 Clinical note Branched-chain aminoaciduria Disease 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 branchedchain 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.

43 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

44 Tetrahydrobiopterin as a cofactor of hydroxylases Dihydrobiopterin

45 Clinical note 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.

46 Tryptophan catabolism Tryptophan has complex catabolic pathway: 1. the indol ring is ketogenic 2. the side chain alanin gluconeogenesis Xanthurenic acid is excrete in the urine. Nicotinamide NAD and NADP.

47 Enzymes which metabolised amino acides containe vitamines as cofactors THIAMINE B 1 (thiamine diphosphate) oxidative decarboxylation of a-ketoacids RIBOFLAVIN B 2 (flavin mononucleotide FMN, flavin adenine dinucleotide FAD) oxidses of a-amino acids NIACIN B 3 nicotinic acid (nikotinamide adenine dinucleotide NAD + nikotinamide adenine dinukleotide phosphate NADP + ) dehydrogenases, reductase PYRIDOXIN B 6 (pyridoxalphosphate) transamination reaction and decarboxylation FOLIC ACID (tetrahydropholate) Meny enzymes of amino acid metabolism

48 Pictures were taken from textbooks: Marks Basic Medical Biochemistry A Clinical Approach. Four edition M. Lieberman, A.D. Marks ed., Essentials of Medical Biochemistry With Clinical Cases. First edition. N.V. Bhagavan, Chung-Eun Ha ed., 2011.