Amino Acids & Proteins

Amino Acids Amino acids are organic compounds containing both amino group & carboxyl group, those occurring in human proteins are L α-amino acids having the formula RCH(NH2)COOH & with the exception of glycine all amino acids are asymmetric .

D-Amino acids that occur naturally are free D-serine& D- aspartate which present in the in brain tissue.

Classification of Amino acids Over 300 amino acids occur naturally, however; only 20 of them share in the formation of proteins. Classification of amino acids either according to their R-group or to their nutritional importance.

1-Classification according to R-group(side chain).

1

2

2-Classification according to their nutritional importance All 20 amino acids present in proteins are biologically essential but not all of them are nutritionally essential, therefore, they are classified into:

3 A-Nutritionally essential amino acids :they should be supplement in diet because human body can not synthesize it , there deficiencies lead to kwashiorkor (protein malnutrition) & marasmus (protein & energy malnutrition). B-Nutritionally nonessential amino acids : human body can synthesize it , therefore , it’s not essential to be supplied in the diet.

R-group It determines the property of amino acid in peptide formation as: 1-Since glycine is smallest amino acid, therefore, it can accommodate in places inaccessible to other amino acids in the formation of peptide.

4 2-Hydrophobic R-group of alanine, valine, leucine, Isoleucine, & aromatic R-groups of phenylalanine , tyrosine & tryptophan make them in the interior of protein. 3-The charged R-group of basic & acidic amino acids stabilize protein conformation through ionic interaction or salt-bridge.

In Vivo Synthesis of Nutritionally Nonessential Amino Acids

Glutamate & Glutamine: Reductive amination of α- ketoglutarate is catalyzed by glutamate dehydrogenase enzyme forming glutamate, this reaction is reversible. Amidation of glutamate to glutamine is catalyzed by glutamine synthetase enzyme, this irreversible reaction requires ATP.

Alanine: By amination of pyruvate forming alanine, the amino donor may be glutamate or aspartate, this reversible transamination reaction is catalyzed by transaminase enzyme.

Aspartate & Asparagine: By amination of oxaloacetate forming aspartate, the amino donor may be alanine; this

5 reversible transamination reaction is catalyzed by transaminase enzyme. Amidation of aspartate to asparagine is catalyzed by asparagine synthetase enzyme, this irreversible reaction requires ATP and it’s resemble amidation of glutamate , however , glutamine rather than ammonium ion, provides the nitrogen for this amidation.

Serine:Formed from glycolytic intermediate 3-phosphoglycerate

Glycine : Formed from glyoxylate , glutamate or alanine by a reversible reaction catalyzed by glycine aminotransferases ,it can produce also from choline & serine.

Proline: Formed from glutamate.

Cysteine: Formed from methionine as discussed later.

Tyrosine: phenylalanine converted to tyrosine by irreversible reaction catalyzed by phenylalanine hydroxylase enzyme; since this reaction is irreversible ,therefore, dietary tyrosine can not replace phenylalanine.

6

Hydroxyproline & Hydroxylysine : They are important in posttranslational processing of collagen & not necessary for protein synthesis .Formed by hydroxylation of proline & lysine in a reaction catalyzed by prolyl hydroxylase &lysyl- hydroxylase enzymes respectively, it require vitamin C as cofactor , therefore , deficiency of vitamin C lead to scurvy.

Proteins Formation of peptides Link of amino acids lead to the formation of peptides. -If the chain formed from up to five amino acids the structure is called oligopeptide or called dipeptide, tripeptide, tetrapeptide & so on if consist from 2,3,4 amino acids respectively. -If the chain formed from 6-30 amino acids the structure is called polypeptide. -If the chain formed from larger than 30 amino acids the structure is called protein.

Classifications of Proteins ►I-Initial classification according to their properties as: 1-Solubility : 2-Shape :

7 A-Globular proteins: roughly spherical or ovoid in shape with their axial ratios (ratio of their longest to shortest dimensions) of not more than 3 with tendency to be soluble in physiological fluids(because it is compact with hydrophilic R-group face outward ), it represent nearly all proteins of interest in clinical chemistry like hemoglobin , enzymes & plasma protein except fibrinogen. B-Fibrous proteins: their axial ratios 10 or more with tendency to be insoluble in physiological fluids e.g fibrinogen. 3-Presence of nonproteins groups (prosthetic groups): these prosthetic groups are bound to protein forming conjugated proteins , the conjugated proteins without prosthetic groups are called apoproteins. prosthetic group + apoprotein = conjugated protein example: A-lipid + protein = lipoproteins B- CHO+protein=glycoprotein(if CHO 5-15%)& mucoprotein (if CHO 15-75%) C-metal ion+ protein = metalloprotein as hemoglobin . ►II-Nowadays with the development of determination of amino acids sequences of proteins more precise classification depend on homology of sequence & structure of amino acids in protein , however , early classification still in use.

Orders of Protein Structure Conformation of proteins into the following orders of structure is necessary to maintain their function & physical property. 1-Primary structure 2-Secondary structure 3-Tertiary structure 4-Quaternary structure

Primary structure Represent the sequence of amino acids in polypeptide chain , its genetically determined, formed by peptide-bonds that linked amino acids together by connecting amino group of one amino

8 acid with a carboxyl group of another leaving only one free amino group & carboxyl group at both extreme of protein, the amino acids present in peptide are called aminoacyl residues & are named by replacing the ate or ine suffixes of free amino acids with yl e.g..(alanyl,aspartyl,glutamine) the ine ending of glutamine indicates that it is carboxyl group is not involved in the peptide bond formation (extreme of protein).

Secondary structure Generally peptide bonds of protein restrict this secondary conformation , however , free rotation is possible only in a bond that link : a) α-carbon to the carbonyl carbon b) α-carbon to the nitrogen This secondary structure has the following conformations: 1-Alpha Helix 2- Beta Sheet 3-Loops & Bends 1-Alpha Helix: look like cylinder with R-group of each aminoacyl residue face outward.

9 2-The Beta Sheet: look like zigzag or pleated pattern with R- groups of adjacent residues point in opposite directions , unlike the compact backbone of α-helix the backbone of Beta Sheet is highly extended , most Beta Sheet are twisted & not perfectly flat forming the core of globular proteins.

3-Loops & Bends : It refer to the short segments of amino acids that joint two units of secondary structure & it represent the key of biological role of that protein.

Tertiary structure Secondary structural features as alpha-helix , beta-sheet , loops & bends are assemble to form domain which represent a section of protein structure sufficient to perform a particular physical or chemical task. Some polypeptides are small consist from a single domain as myoglobin while other are large consist from many domains. The tertiary structure represents how secondary structural features are assemble to form domain & how these domains relate spatially to one another.

11 Quaternary structure Some proteins are assembled from more than one polypeptide or protomer. Quaternary structure defines the polypeptide composition of protomer & for those protein containing more than one protomer the spatial relationships between these protomers . Protein with a single protomer called monomeric protein. Protein with two protomers called dimeric protein , these two protomers either identical ((homodimeric)) or not identical ((heterodimeric)). Greek letters ((α,β,γ… )) are used to distinguish different protomers in a single protein.

Pathology of Altered Protein Conformation 1-Creutzfelt-jakob disease : Its fatal neurodegenerative disease resulting from alteration of protein conformation in neural cells , the pathogenesis of disease is still not clear probably due to chromosomal changes ((chromosome 20)) . 2-Alzheimer's disease: It characterize by senile plaques in the brain with alteration in neuron protein conformation, the pathogenesis of disease is still not clear. 3-Thalassemia: caused by genetic defects that impair the synthesis of one of the polypeptide subunits (α or β) of hemoglobin.

Biological Functions of Proteins 1-Enzymes: catalyze biochemical reactions. 2-Peptide hormones : regulate metabolism. 3-Antibodies & Complements : defense mechanism . 4-Albumin : maintain oncotic pressure of plasma. 5-Many plasma proteins like albumin transport vitamins, metals, hormones & drugs in the blood often serving as reservoirs . 6-Hemoglobin : carry oxygen. 7-Coagulation factors : affect haemostasis. 8-Structural proteins as keratin & collagen. 9-Contractile proteins as myosin. 10-Storage proteins as ferritin that store iron.

11 Digestion & Absorption of Proteins Few bonds are accessible to the proteolytic digestive enzymes (proteases) that catalyze hydrolysis of peptide bonds, without prior denaturation of dietary proteins (by heat in cooking & by the action of gastric acid). There are two main classes of proteases enzymes which are:- A-Endopeptidases:- hydrolyze peptide bonds between specific amino acids throughout the molecule. They are the first enzymes to act, yielding a larger number of smaller fragments. E.g.; of endopeptidases are pepsin in the gastric juice; trypsin, chymotrypsin & elastase secreted into the small intestine by the pancreas. B-Exopeptidases catalyze the hydrolysis of peptide bonds from the ends of peptides; they include 1-Carboxypeptidases: secreted in the pancreatic juice, release amino acids from the free carboxyl terminal. 2- Aminopeptidases: secreted by the intestinal mucosal cells, release amino acids from the amino terminal. 3-Dipeptidases :secreted in the brush border of intestinal mucosal cells catalyze the hydrolysis of dipeptides, which are not substrates for amino- and carboxypeptidases. The end product of protein digestion is amino acids which are absorbed through the portal vein to reach the liver.

Catabolism of Proteins The susceptibility of protein to degradation is expressed by its half-life((T1/2)) which define as a time required to lower protein concentration to half of its initial value, each day human body turn over 1-2% of their total body protein specially muscle protein. Intracellular proteins are degraded to their amino acids by proteases enzymes which include both endopeptidases & exopeptidases ((as in digestive system)). About 75% of liberated amino acids are reutilized for new protein synthesis & because there is no storage for amino acids the rest are rapidly degraded by:

12 1-Catabolism of amino acid nitrogen. 2-Catabolism of the carbon skeleton of amino acid.

Catabolism of Amino Acid Nitrogen The maintenance of steady-state concentrations of circulating plasma amino acids between meals depends on the net balance between release from endogenous protein stores & utilization by various tissues. Muscle generates over half of the total body pool of free amino acids & liver is the site of the urea cycle enzymes necessary for disposal of excess nitrogen. Muscle & liver thus play major roles in maintaining circulating amino acid levels.

In normal adults nitrogen input is equal nitrogen output. Positive nitrogen balance mean that nitrogen input is higher than nitrogen output as in growth & pregnancy. Negative nitrogen balance mean that nitrogen input is lower than nitrogen output as in advanced cancer, following surgery, marasmus & kwashiorkor. The sources of ammonia in the body are: 1- By enteric bacteria which absorbed into portal venous blood. 2- From amino acid catabolism. Ammonia is highly toxic to body specially to C.N.S. because it react with α-ketoglutarate to form glutamate so depletion of α- ketoglutarate impair function of TCA cycle in the neuron , therefore, ammonia should removed rapidly from the circulation by the liver to be converted into urea so only traces of ammonia ((10-20µg/dl)) are present in the peripheral blood. Ammonia intoxication occur in sever liver impairment & it presented clinically as tremor , slurred speech , blurred vision , abnormal behavior , coma & death.

Biosynthesis of Urea Urea biosynthesis occurs in four stages: 1-Transamination. 2- Oxidative deamination of glutamate.

13 3-Ammonia transport. 4-Reactions of urea cycle.

Transamination It involves transfers of α-amino nitrogen to α-ketoglutarate forming glutamate. All protein amino acids except lysine, threonine, proline & hydroxyproline participate in transamination. The reaction is reversible so it participates in amino acid synthesis as well & it needs the coenzyme pyridoxal phosphate which serves as a carrier of amino groups. Following removal of amino nitrogen by transamination, the remaining carbon skeleton of amino acid is catabolized as discuss later. Generally transamination interconvert pairs of α-amino acids & α-ketoacids catalyzed by aminotransferase (transaminase) enzyme, each aminotransferase is specific for one pair of

substrates but not specific for other pair , in this reaction α- ketoglutarate & glutamate are not specific substrates so that the

14 amino nitrogen of amino acids that undergo transamination was concentrated into glutamate alone & this is important because L- glutamate is the only amino acid that undergoes oxidative deamination at appreciable rate & so forming urea. e.g. of aminotransferase: AST(SGOT) & ALT(SGPT).

Oxidative Deamination of Glutamate Release of α-amino nitrogen of L-glutamate as ammonia is catalyzed by hepatic L-glutamate dehydrogenase enzyme which can use either NAD+ or NADP+, this reaction is reversible so it participates in amino acid synthesis as well. Conversion of α-amino nitrogen of amino acid to ammonia by the concerted action of aminotransferase & L-glutamate dehydrogenase enzymes is often termed ((transdeamination)).

Other mechanisms: 1))) - Glucose-alanine cycle: amino group transported from the muscle to the liver in the form of alanine & in the liver the amino group is converted to urea by the urea cycle.

2))) - L-amino acids oxidases enzymes of the liver & kidney convert amino acid to keto acid with release of ammonium ion , the physiological role of this reaction is still uncertain. 3))) - Formation of glutamine from glutamate is catalyzed by glutamine synthetase enzyme, one function of glutamine is to sequester ammonia in a nontoxic form. Hydrolysis release of the

15 amide nitrogen of glutamine as ammonia is catalyzed by glutaminase enzyme in an irreversible reaction, therefore, the concerted action of glutamine synthetase & glutaminase is to catalyze the interconversion of free ammonium ion & glutamine. Analogous reaction of asparagine is catalyzed by asparagine synthetase & asparaginase enzymes. A rare deficiency in neonate glutamine synthetase results in severe brain damage, multi-organ failure & death.

Ammonia Transport Because of high ammonia toxicity, these produced from amino acid catabolism are rapidly removed by the liver while ammonia produced by enteric bacteria is absorbed into portal venous blood to the liver. Liver convert ammonia to urea which is less toxic by urea cycle, the produced urea is removed from the body mainly by the kidney.

Reactions of urea cycle It consists of ((5)) reaction steps:- (Step-1)Carbamoyl phosphate synthesis. (Step-2) Citrulline synthesis. (Step-3) Argininosuccinate synthesis. (Step-4) Argininosuccinate cleavage. (Step-5) Urea release.

(Step-1)Carbamoyl phosphate synthesis: Its the rate limiting reaction of urea cycle, it involve the condensation of CO2 + ammonia + ATP to form carbamoyl phosphate, this irreversible reaction occurs inside the mitochondria & is catalyzed by mitochondrial carbamoyl phosphate synthase I enzyme which require N-acetylglutamate as enzyme activator, this reaction require 2 moles of ATP ((one ATP for energy while second ATP provide phosphate for synthesis of carbamoyl phosphate )).

16

(Step-2) Citrulline synthesis: It involve the transfer of carbamoyl group ((H2N-C= O)) of carbamoyl phosphate to ornithine forming citrulline & phosphate , this irreversible reaction is catalyzed by Ornithine transcarbamoylase enzyme & it occurs in the mitochondria , however , both the formation of ornithine & subsequent metabolism of citrulline take place in the cytoplasm.

(Step-3) Argininosuccinate synthesis: It involve the link of aspartate & citrulline via the amino group of aspartate to form argininosuccinate ,this irreversible reaction is catalyzed by argininosuccinate synthase enzyme & it require one mole of ATP , this reaction provides the second nitrogen of urea ((from aspartate))& it occurs inside the cytoplasm.

17

(Step-4) Argininosuccinate cleavage: This cleavage form arginine (retain nitrogen) + fumarate, this irreversible reaction occurs inside the cytoplasm & is catalyzed by argininosuccinase enzyme.

(Step-5) Urea release: Hydrolytic cleavage of arginine is catalyzed by arginase enzyme producing urea + ornithine, this irreversible reaction occurs inside the cytoplasm. Ornithine reenters liver mitochondria for additional rounds of urea biosynthesis.

Summary of urea cycle 1- Synthesis of 1 mole of urea requires 3 moles of ATP + 1 mole of ammonium ion + 1 mole of aspartate + 1 mole of Co2 . 2- It consist of 5 irreversible reactions each reaction is catalyzed by a single enzyme, therefore, 5 enzymes was involved in

18 addition the six participating amino acids N-acetylglutamate functions as an enzyme activator. 3- There is no net loss or gain of major metabolites ornithine , citrulline , argininosuccinate & arginine ((cyclic process)) , however, ammonium ion , CO2 , ATP & aspartate are consumed. 4- Some reactions occur in the mitochondria while others occur in the cytoplasm. 5-Major changes of diet can increase the concentrations of individuals urea cycle enzymes 10-20 folds as in starvation which elevates the enzyme levels to cope with the increased production of ammonia that accompanies protein degradation.

19 Metabolic disorders of urea cycle They are extremely rare , however, they have the following principles: 1))- Similar clinical picture {ammonia intoxication}, however , clinical presentation is more severe if block occur at reaction 1 or 2 because once citrulline is synthesized there is some link between ammonia & carbon so serve as alternative carrier of excess nitrogen . 2))- Pathogenesis is mainly due to defect in the specific enzyme. 3))- Accumulation of intermediates & of ancillary products that accumulate prior to metabolic block give an idea about the impaired reaction . 4))- Precise diagnosis require quantitative assay of the activity of the defective enzyme. 5))- Diet therapy includes the ingestion of frequent small low protein diet to avoid sudden increase of blood ammonia levels. 6))- Rational therapy must be based on the understanding of biochemical reaction. These metabolic disorders include: Hyperammonemia Type 1: deficiency of the enzyme carbamoyl phosphate synthase I. Hyperammonemia Type 2: deficiency of the enzyme Ornithine transcarbamoylase . Patient has a high level of glutamine in the blood , CSF & urine as response to elevated level of tissue ammonia. N-Acetylglutamate Synthase Enzyme Deficiency: Deficiency of this enzyme that is responsible for the synthesis of N-Acetylglutamate which is essential for carbamoyl phosphate synthetase I activity result in clinical & biochemical features indistinguishable from those arising from a defect in carbamoyl phosphate synthetase I(Hyperammonemia Type 1), however it may respond to N-acetylglutamate adminstration. Citrullinemia: deficiency or defective activity of argininosuccinate synthase enzyme. Patient has a high level of citrulline in the blood , CSF & urine.

21 Argininosuccinicaciduria: deficiency of argininosuccinase enzyme. Patient has a high level of argininosuccinate in the blood , CSF & urine . Hyperargininemia: deficiency of arginase enzyme. Patient have a high level of arginine in the blood, CSF & their urine pattern are resemble that of lysine-cystinuria due to competition of arginine with lysine & cystine for reabsorption site of renal tubules. HHH syndrome: defect in the mitochondrial membrane ornithine transporter result in the development of hyperornithinemia, hyperammonemia & homocitrullinuria syndrome (HHH syndrome), The failure to import cytosolic ornithine into the mitochondrial matrix renders the urea cycle inoperable, with consequent hyperammonemia & the accompanying accumulation of cytosolic ornithine results in hyperornithinemia. In the absence of its normal acceptor (ornithine), mitochondrial carbamoyl phosphate carbamoylates lysine to homocitrulline with a resulting homocitrullinuria.

21 Catabolism of Carbon Skeletons of Amino Acids Transamination initiate amino acid catabolism ((except proline, hydroxyproline, threonine, lysine)) , the residual hydrocarbon skeleton is then degraded to intermediates . Metabolic defects (Inborn errors of metabolism) associated with these process many of them require prenatal (measure enzyme activity in cultured amniotic fluid cells)or early postnatal diagnosis & timely initiation of treatment to avoid their sequel. Many of these disorders characterized by aminoaciduria. Aminoaciduria: may be primary or secondary A-Primary: due to inborn error of metabolism ,therefore, affect specific amino acid .It is either overflow or renal type 1-Overflow: defect in the pathway of amino acid catabolism , characterize by a high level of amino acid or their metabolites in blood & urine. 2-Renal :defect in renal tubular reabsorption of specific amino acid , characterize by a high level of amino acid in urine only(not in blood). B-Secondary: due to diseases affecting many amino acids simultaneously. It is either overflow or renal type 1-Overflow: as in acute hepatic failure 2-Renal : as in diseases affecting proximal renal tubules where reabsorption of amino acids occurs. The catabolism of carbon skeleton of amino acids is subdivided into:- I-Amino Acids Form Pyruvate or Intermediate of Krebs cycle: These amino acids are a source of energy (through enterance to Krebs cycle) or they may be used for glucose synthesis by gluconeogenesis, therefore, called glucogenic amino acids. II-Amino Acids Form Acetyl-CoA: These amino acids are a source of energy(through enterance of acetyl-CoA to Krebs cycle) in addition to the formation of ketone bodies from acetyl- CoA, therefore, called ketogenic amino acids. Note: Some amino acids have both ketogenic & glucogenic properties.

22 I-Amino Acids Form Pyruvate or Intermediate of Krebs cycle

*Asparagine , Aspartate , Glutamine & Glutamate: Asparagine & aspartate form oxaloacetate. Analogous reaction converts glutamine & glutamate to α-ketoglutarate . No metabolic defect is associated with these amino acids metabolism.

*Proline: Proline forms finally α-ketoglutarate as follow:

Metabolic defect 1-Type I hyperprolinemia: metabolic block at proline dehydrogenase enzyme ((hydroxyproline not affected)). 2-Type II hyperprolinemia: metabolic block at glutamate- semialdehyde dehydrogenase enzyme which share also in hydroxyproline catabolism, therefore, both proline & hydroxyproline are affected.

*Arginine: Arginine is converted to ornithine in irreversible reaction catalyzed by arginase enzyme, then ornithine is converted to L-Glutamate-γ-semialdehyde by reversible reaction catalyzed by ornithine aminotransferase enzyme, finally L- Glutamate-γ-semialdehyde is converted to α-ketoglutarate.

23

Metabolic defect 1-Hyperargininemia: deficiency of arginase enzyme. 2-Mutation of ornithine aminotransferase: elevate plasma & urinary ornithine causing retinal atrophy, treatment by restricting dietary arginine. 3-HHH syndrome: defective of ornithine mitochondrial transport.

*Histidine: Catabolism of histidine through many steps ends in formation of α-ketoglutarate: Metabolic defect Histidinemia: Benign disorders due to impaired histidase enzyme which is the first enzyme involved in histidine catabolism.

*Glycine: Two pathways: 1-The glycine cleavage complex of liver mitochondria split glycine to CO2 , NH4 & conversion of tetrahydrofolate into 5,10- methylene tetrahydrofolate . 2-Reversible transamination reaction catalyzed by glycine aminotransferase enzyme lead to deamination of glycine forming glyoxylate which change to pyruvate. Metabolic defect 1-Glycinuria: defective renal tubular reabsorption of glycine.

24 2-Primary hyperoxaluria: failure to catabolize glyoxylate, subsequent oxidation of accumulated glyoxylate to oxalate lead to hyperoxaluria manifested clinically as urolithiasis & nephrocalcinosis which may lead to hypertension & renal failure.

*Serine: There is interconversion between serine & glycine by reversible reaction catalyzed by serine hydroxymethyltransferase enzyme, therefore, serine catabolism merges with that of glycine.

*Alanine: Transamination of alanine form pyruvate, no metabolic defect of alanine catabolism.

*Cysteine: Initially one molecule of cystine is reduced to two molecules of cysteine by cystine reductase enzyme.

Two different pathways that convert cysteine to pyruvate. 1)- Cysteine sulfinate pathway. 2)- 3-mercaptopyruvate pathway. Metabolic defect 1-Cystine-lysinuria(Cystinuria): defect in renal tubular reabsorption of cystine , lysine , arginine & ornithine , therefore

25 , they excreted in urine , however , the condition is benign apart from cystine calculi , therapy by increase water intake , increase cystine solubility by maintaining an alkaline urine . 2-Cystinosis(cystine storage disease): defect of cystine transport with deposition of cystine crystals in tissues &early mortality of acute renal failure.

*Threonine: Threonine aldolase enzyme cleaves threonine into glycine & acetaldehyde. Glycine is catabolized into pyruvate while acetaldehyde is converted into acetyl-CoA so that threonine is an example of both ketogenic & glucogenic amino acid.

*Hydroxyproline: Catabolism ends in pyruvate formation:

Metabolic defect 1-Hyperhydroxyprolinemia: block at hydroxyproline dehydrogenase enzyme , benign condition , no associated impairment of proline catabolism. 2- Type II hyperprolinemia: block at γ-hydroxy- glutamate- γ-semialdehyde dehydrogenase enzyme whish share also in proline catabolism, therefore, both proline & hydroxyproline are affected.

26 *Methionine: Catabolism of methionine end in the formation of succinyl-CoA.

Methionine Homocysteine

Cystathionine synthase Homocysteine + Serine Cystathionine

Cystathionine Cysteine + Succinyl-CoA(many steps)

Metabolic defect Homocystinuria: metabolic defect in cystathionine synthase enzyme, therefore, homocysteine & methionine concentrations are elevated in the blood & can be detected in urine while cysteine concentration is reduced in the blood. The most serious sequel of homocystinuria is thrombosis which; therefore, neonatal screening by measuring plasma level of methionine is necessary. Treatment by dietary restriction of methionine with cysteine supplement.

*Valine: Discussed later with branched chain amino acids

27 II-Amino Acids Form Acetyl-CoA

*Tyrosine: End product of tyrosine catabolism is acetyl-CoA:

Metabolic defect 1-Since ascorbate is needed for p-hydroxyphenylpyruvate hydroxylase enzyme, therefore , it is deficiency lead to excretion of incomplete products of tyrosine catabolism. 2-Type I tyrosinemia (tyrosinosis): Defect in fumarylacetoacetate hydrolase enzyme [[ 4 in figure]] , untreated cases lead to death from liver failure & generalized renal tubular failure, treatment by low tyrosine & phenylalanine diet. 3-Type I I tyrosinemia (Richner-Hanhart syndrome): Defect in tyrosine transaminase enzyme[[ 1 in figure]] , therapy by low tyrosine & phenylalanine diet. 4-Neonatal tyrosinemia: Defect in p-hydroxyphenylpyruvate hydroxylase enzyme [[ 2 in figure]] , it is transient condition occur in small for date (SFD) & in premature baby ,therapy by low tyrosine & phenylalanine diet. 5-Alkaptonuria: metabolic defect in homogentisate oxidase enzyme [[ 3 in figure]] , during infancy & childhood the patient

28 have dark urine due to oxidation of excreted homogentisate in urine . Later in life patient have arthritis & connective tissue pigmentation first appear in the ears(ochronosis) due to oxidation of homogentisate which polymerized & binds to connective tissue. Treatment :restriction of tyrosine & its precursor phenylalanine.

*Phenylalanine: Catabolism of phenylalanine as follow: A-Major pathway: Phenylalanine is converted to tyrosine which metabolized into acetyl -CoA as follow:

B-Minor(Alternative )pathway: occur in normal liver as follow :

29 Metabolic defect Hyperphenylalaninemias : they are group of disorders (Types I-V) resulting from impaired conversion of phenylalanine to tyrosine(major pathway), however , the most common one is Type I (classic phenylketonuria or PKU) where metabolic defect at phenylalanine hydroxylase I , patient have a high blood level of phenylalanine with stimulation of minor pathway lead to increase renal excretion of phenylpyruvate & it is metabolites which react with Fecl3 as ketone hence the name PKU. Routine neonatal screening is required to avoid mental retardation by doing : 1-Fecl3 urine test to detect phenylpyruvate 10-14 days after birth. 2-More reliable by measuring phenylalanine in blood 3-4 days after birth or measurement of enzyme activity. Treatment by low phenylalanine diet.

*Lysine: The first six reactions of L-lysine catabolism in human liver form crotonyl-CoA, which is then degraded to acetyl-CoA & CO2. Metabolic defect of any enzyme involved in lysine catabolism lead to hyperlysinemia.

*Tryptophan: It is degraded via the kynurenine-anthranilate pathway that includes many reaction steps lead finally to the formation of acetyl-CoA. Metabolic defect 1-Since kynurenine-anthranilate pathway require pyridoxal phosphate(PLP) , therefore , in vitamin B6 deficiency there is impairment of this pathway with consequent accumulation of intermediate which converted into xanthurenate, therefore, urinary excretion of xanthurenate in response to tryptophan load is a diagnostic of vitamin B6 deficiency. 2-Niacin is synthesized from tryptophan , therefore , in Hartnup disease ((impairment of intestinal & renal transport of tryptophan)) there is niacin deficiency lead to pellagra.

31 *Branched Chain Amino Acids: They include Leucine , Isoleucine & valine , their catabolism as follow : Reaction 1(Transamination) : Convert the branched chain amino acids into α-ketoacids. Reaction 2(Oxidative decarboxylation) : Catalyzed by a multienzyme complex called branched chain α-ketoacid decarboxylase complex Reaction 3(Dehydrogenation) :The products of this reaction enter multiple steps reaction end finally in the formation of : *Acetyl-CoA in leucine & isoleucine. *Succinyl-CoA in valine. Metabolic defect *Maple syrup urine disease (branched-chain ketonuria): metabolic defect in branched chain α-ketoacid decarboxylase complex result in the accumulation of branched chain amino acids & their corresponding α-ketoacids in blood & urine lead to acute ketoacidosis & later on mental retardation . Neonatal diagnosis is necessary by: 1-Earliest diagnosis by measuring enzyme activity. 2- Estimating a high level of branched chain amino acids in blood & urine. 3- Presence of elevated level of their α-ketoacids in urine using 2,4-Dinitrophenylhydrazine test which show yellow precipitate of hydrazone . Treatment by dietary restriction of leucine , isoleucine & valine.

31 Conversion of amino acids to specialized products Amino acids can be converted to many specialized products as: Glycine: Importance as follow: 1-Metabolites & many drugs are excreted as water-soluble glycine conjugates. 2-Participate in biosynthesis of creatine. 3 -Participate in biosynthesis of heme. 4 -Participate in biosynthesis of purines. Phosphorylated Serine , Threonine & Tyrosine: Importance in regulation of certain enzymes of lipid & CHO metabolism. Methionine: Importance of S-Adenosylmethionine in: 1-Principle source of methyl groups in the body so share in creatine synthesis. 2-Participate in biosynthesis of polyamines. What are polyamines ? : include spermine & spermidine, have a role in: 1-Function in cell proliferation & growth. 2-Growth factors for cultured mammalian cells. 3-Stabilize intact cells. Cysteine: Importance in : 1-Participate in coenzyme A synthesis. 2 Participate in taurine synthesis that conjucates with bile acids . Histidine: Importance in histamine synthesis. Arginine: Importance in : 1- Share in creatine synthesis. 2- Via ornithine share in polyamines synthesis. 3-Arginine is a precursor for intracellular signaling molecule nitric oxide (NO) that serves as neurotransmitter , smooth muscle relaxant & vasodilator. Tryptophan: Importance in: 1-Niacin synthesis, therefore tryptophan deficiency leads to pellagra. 2-Serotonin synthesis: this is done in argentaffin cells which present mainly in the ileum & appendix,less extent in pancreas , rectum & stomach . The serotonin synthesis is as follow:

32 Tryptophan

Tryptophan 5-hydroxylase

5-Hydroxytryptophan(5HTP)

Aromatic amino acid decarboxylase

5-Hydroxytryptamine(5HT,Serotonin)

Monoamine oxidase

5-Hydroxyindole acetic acid(5HIAA) ---urine

Argentaffin cells also produce a peptide called substance-P Carcinoid tumour(Argentaffinoma) is caused by tumour of argentaffin cells produce excess amount of 5HTP , 5HT & their urinary metabolites 5HIAA. Patient complain mainly from sever diarrhea , flushing , bronchospasm , these symptoms mainly due to excess substance-P rather than excess serotonin , patient also may complain from pellagra because tryptophan is diverted from niacin synthesis to serotonin synthesis. Tyrosine: Importance in : 1- Precursor of catecholamines (Dopamine, Norepinephrine & Epinephrine). 2-Formation of melanin. 3-Precursor of T3 & T4 hormones produced by thyroid gland. Creatinine: formed in muscle from creatine phosphate :

Therefore, arginine , glycine & methionine share in creatinine biosynthesis which is related to muscle mass. Glutamate: Decarboxylation of L-glutamate leads to formation of γ-Aminobutyric acid (GABA) which is inhibitory neurotransmitter in the brain.

33