Keshava Pavan K, medical student, KMC, Mangalore, India AMINO ACIDS AND PROTEINSImportant points → All amino acids in body are l- amino acids → Dextro/laevo form of amino acid is decided by –NH2 group → There are amino acids other than the 20 but they are not found in proteins (as they are not coded) → Above mentioned amino acids come under DERIVED AMINO ACIDS → Hydroxyproline, hydroxylysine, γ-carboxyglutamic acid, N- Formyl methionine are derived amino acids found in proteins → L-ornithine, citrulline, GABA, homoserine, homocysteine, histamine are derived amino acids not found in proteins → Non- alpha amino acids: β-alanine, GABA, δ-aminolevulinic acid, taurine → D- amino acids: Actinomysin D, gramicidin, polymyxin, valinomysin → Proteins in lipid environment have non-polar amino acids → At isoelectric pH, amino acids have least mobility, minimum solubility & least buffering action → Histidine is a very good buffer in the body as its pI (7.6) is close to blood pH → Histidine is found mainly in hemoglobin → A biologically active protein has atleast a 3D structure → Collagen is most abundant protein in human body → STRUCTURES OF COLLAGEN IN DIFFERENT PLACES Vitrious humour- dispersed as gel to stiffen structure Tendons- bundled in tight parallel fibers to provide tensile strength Cornea- stacked to transmit light with minimum scattering Bone- → Parallel arrangement is quarter staggered
→ Osteogenesis imperfecta: replacement of glycine→ GLUTATHIONE Tripeptide Maintains RBC membrane structure integrity Protects Hb from oxidation by H2O2 detoxification→ Substance- P Decapeptide Neurotransmitter→ Denaturating agents Physical: heat, violent shaking, X-rays, UV rays Chemical: acid, alkali, organic solvents, heavy metal salts, urea, salicylate→ Denaturation of ribonuclease by urea is reversible→ Denatured protein cannot be crystallised→ Complete proteins: egg albumin, milk proteins→ Partially incomplete proteins: rice & wheat proteins (no Lys, Thr)→ Incomplete proteins: gelatin (Trp), zein (Trp, Lys)→ TDM : Therapeutic Drug Monitering→ Functional classification Structural – keratin, collagen, elastin Catalytic – enzymes Transport – Hb, albumin, transferrin Hormonal – insulin, GH Contractile – actin, myosin Storage – ferritin, myoglobin
Genetic – nucleoprotein Immune – immunoglobulins (Igs) Receptor – for hormones, viruses etc. Plasma proteins – albumin, globulin, fibrinogen, prothrombin→ Total plasma protein concentration : 6-8 g/dL→ Albumin – 3.5 – 5.5 g/dL→ Globulin – 1.8-3.6 g/dL→ Fibrinogen – 0.2- 0.4 g/dL→ Types of globulins→ α1 globulin Retinal binding protein (RBP) (binds with Vit A & transports it) α1 fetoprotein α1 antitrypsin apolipoprotein A→ α2 globulin ceruloplasmin transcortin haptoglobin→ β globulin β-hemopexin transferrin C-reactive protein→ γ globulin immunoglobulins→ Albumin 50-60% of plasma proteins
Simple, globular protein Synthetized in hepatocytes Half life of 15- 20 days Undergoes degradation- pinocytosis→ Functions: Osmotic pressure regulation Blood volume & blood pressure regulation Transportation (of mainly bilirubin, fatty acid, Ca2+, Cu2+, drugs like aspirin, sulphonamides, steroid hormones) Nutritive→ Disorders: Hypoalbuminemia: • Malnutrition • Malabsorption • Liver cirrhosis & other liver disorders • Excess loss during nephrotic syndrome and other kidney diseases, surgery & haemorrhage • Overhydration/hemodilution Analbuminemia: • Genetic • Types are homozygous & heterozygous→ Pre albumin – two parts: thyroxine binding pre albumin binds thyroxine; retinal binding pre albumin binds retinal – theses two are found in 1:1 ratio and are together called transthyretin→ Acute phase proteins (APP) Plasma proteins that change in concentration due to acute phase reactions(APR). 2 types – positive & negative
+ increases during APR. e.g,. α1 antitrypsin, haptoglobin, ceruloplasmin, CRP - decreses during APR. e.g,. pre-albumin, albumin, transferrin→ Causes for acute phase reactions Injury/infection/cancer Inflammation Positive response Negative response ↑cytokines ↑vasoactive substances ↑interleukins alter vascular permiability ↑positive APP movement of plasma proteins from plasma to ECF→ α1 antitrypsin prevents elastase from degrading elastin in lungs.→ If α1 antitrypsin is deficient – exposed to smoke – met-sulphoxide formed – emphysema (therefore more in smokers)→ C Reactive protein – binds to C-polysaccharide present on pneumococci→ Ceruloplasmin Glycoprotein Binds to 6 molecules of copper
Binds to 90% of Cu2+ in blood circulation Blue coloured Ferrous to ferric Deficiency- WILSON’S DISEASE (hepato-lenticular degeneration –refer MINERALS – COPPER) Normal concentration- 25 to 50 mg/dL→ IMMUNOGLOBULINS Synthesized by plasma cells Glycoprotein Tetramer of 2 light & 2 heavy chains Constant & variable region Hyper-variable regions – 3 in light chain, 4 in heavy chain.→ Structure – refer diagram (1)→ Hydrolysis at hinge region – refer diagram (2)→ Types of light chains Kappa (K) Lambda (λ)→ Types of heavy chains gamma, γ (IgG) Alpha, α (IgA) Mu, µ (IgM) Delta, δ (IgD) Eta, ε (IgE)→ Individual structures – refer diagram (3)→ Functions of Igs:→ IgG
70% of Igs Secondary immune response Only Ig to cross placental barrier thus providing protection to fetus. Neutralisation of toxins from antigenic cells Enhances activity of complement proteins Prepares cell for phagocytosis (opsonisation)→ IgA 20% of Igs Found in body fluids Mucus & body secretions Surface immunity→ IgM 8 – 10% Primary response Phagocytosis→ IgD Less than 1% Surface receptor Not much known because it is very labile→ IgE 0.004% During allergy, binds to mast cell & basophil, rupturing their membranes releasing histamine. (- hypersensitivity)→ Multiple myeloma Malignant proliferation of plasma cells→ BENCE JONES PROTEIN
Low molecular weight proteins Produced excessively during Multiple myeloma & other disorders Excreted in urine. → Digestion of proteins → In stomach HCl acidic pH helps to activate pepsinogen to pepsin denaturation protection against bacteria pepsin rennin in infants → endopeptidase and exopeptidase activity – refer diagram (4) → in intestine pancreatic enzymes – secretin, cholecystokinin intestinal enzymes – trypsin, chymotrypsin, carboxypeptidase A and B, elastase → Absorption of proteins – refer diagram (5) → Glutathione: γ glutamyl cycle/Meister cycle - refer diagram (6)General reactions of amino acids: → TransaminationAlanine α ketoglutarate PLP ALTPyruvate glutamatePLP – pyridoxal phosphate; ALT – alanine transaminase/SGPT -serum glutamatepyruvate transaminase
Cysteine pyruvate + H 2S + NH4+ Cysteine desulphydratase Glutamine synthetase Glutamate + NH4+ glutamine In liver glutaminase ATP ADP + Pi glutamate + NH 4+ In muscles in liver Glucose pyruvate alanine pyruvate Glu α KG glu α KG α KG + NH4+ UREA → Urea cycle – refer diagram (7) → Disorders related to urea cycle DISORDER DEFECIENCYHyperammonemia type I CPS IHyperammonemia type II OTCCitrullinemia ASSArgininosuccinic aciduria ASLHyperargininemia Arginase → Normal serum urea level: 20 to 40 mg/dL → Increase in serum urea level – uremia → Causes:
Pre-renal • Vomiting, diarrhoea • Excessive degradation of proteins (as in DM) • Major surgery Renal • Nephrotic syndrome • Other kidney diseases Post-renal • Renal stones • Prostate gland enlargement → Decarboxylation reactions PLPHistidine histamine + CO 2 Histidine decarboxylase5-hydroxy tryptophan 5-hydroxy tryptamine/ serotoninGlutamate γ amino butyric acid SIMPLEST AMINO ACID GLYCINE → Glucogenic → Non essential → Synthesis :
FH4 N5 N10 methylene FH4Serine glycine Serine hydroxy methyl transferase Threonine glycine + acetaldehyde Threonine aldolase → Catabolism: NAD+ NADH PLPGlycine CO 2 + NH4+ Glycine FH4 N5N10 methylene FH4 cleavage systemThe reverse reaction is used in synthesis of glycine. Enzyme is then named glycinesynthase.Deficiency of enzymes of glycine cleavage system causes non-ketotic hyperglycemiaGlycine serine pyruvate gluconeogenesis Serine dehydratase
NH4+ MalateGlycine glyoxalate Oxalate Glycine oxidase FormateBlock in the reaction forming formate leads to increased oxalate in urine(hyperoxaluria) causing urolithiasis, nephrocalcinosis. → Biologically important products formed from glycine: Glutathione Creatine → Glutathione: γ - glutamyl cysteinyl glycine ATP ADP +PiGlutamate + cysteine glutamyl cysteine Glutamyl cysteine synthetase ATP Glycine glutathione synthetase ADP +Pi Glutathione → Creatine – refer diagram (8) → Functions of glycine: Synthesis of glutathione, creatine Component of proteins. Eg., in collagen, every third amino acid is glycine (X – Y – Gly)n In heme synthesis – for details refer PORPHYRINS chapter In conjugation
Cholic acid + glycine glycocholic acidChenodeoxy cholic acid + glycine glyco chenodeoxycholic acidBenzoic acid + glycine hippuric acid → Disorders: Non ketotic hyperglycemia Primary hyperoxaluria Glycinuria – due to defective reabsorption in nephron. SULPHUR CONTAINING AMINO ACIDS METHIONINE → Glucogenic → Essential → Functions: Component of proteins and peptides Coded by initiator codon In transmethylation reactions → Synthesis of functional form SAM – refer diagram (9) → N-methylation: SAM SAHGuanidoacetate creatine Guanidoacetate methyl transferaseNorepinephrine epinephrineNicotinamide N-methyl nicotinamide excreted in urine(detoxification/biotransformation reaction)
→ O-methylation: SAM SAH N-acetyl serotonin O-methyl N-acetyl serotonin/melatonin SAM SAH Epinephrine metanephrine Catechol O-methyl transferase → For summary of methionine refer diagram (10) → Deficiency of cobalamin leads to folate trap → Important reactions of methionine: → Trans-sulphuration pathway: Cystathionine synthaseHomocysteine + serine cystathionine PLP H 2O cystathionase H 2O PLP Homoserine + cysteine
→ Glucogenic pathwayHomoserine α keto butyrate propionic acid NH2 CO2 Glucogenic TCA cycle succinyl CoAMethionine α keto γ methiol butyrate α keto butyrate + methyl mercaptan (CH3 – SH) → Inborn errors of methionine metabolism: → Homocystinuria2 homocysteine homocystine excreted in urine Mental retardation Osteoporosis Intravascular clotting Ectopia lentis High methionine in serum Due to deficiency of either cystathionine synthase or methyl transferase → Cystathioninuria Deficiency of cystathionase Mental retardation CYSTEINE → Non essential → Glucogenic → It exists as cystine
Functions: → Component of proteins and peptides -SH group of glyceraldehyde 3 PO4 dehydrogenase is from cysteine Tertiary and quaternary structures of proteins result from disulfide linkages between cysteine residues as in insulin, immunoglobulins. Keratin has high concentration of cysteine Collagen does not have cysteine. → Component of glutathione Glutathione is called a pseudopeptide as peptide bond is between γ-carboxyl group and α-amino group instead of α-carboxyl group. All peculiar properties of glutathione are due to –SH group of cysteine. 2 G-SH G-S-S-G → Thioethanolamine of CoASH Components of coenzyme A are • Pantothenic acid (pantoic acid + β alanine) • β mercapto ethanolamine/thioethanolamine • AMP • Pyrophosphate The thioethanolamine component is derived from cysteine. → DetoxificationBromobenzene mercapturic acid cysteine of glutathione → Formation of taurine Primary bile acids – cholic acid, chenodeoxy cholic acid Conjugated by glycine and taurine Primary conjugated bile acids formed – glycocholic acid, taurocholic acid, glycochenodeoxy cholic acid, taurochenodeoxy cholic acid These combine with sodium or potassium to form bile salts – sodium glycocholate, potassium glycocholate, sodium taurocholate, potassium taurocholate, sodium glycochenodeoxy cholate, potassium
glycochenodeoxy cholate, sodium taurochenodeoxy cholate and potassium taurochenodeoxy cholate. Formation of taurine from cysteine: refer diagram (11)Inborn errors of cysteine metabolism: → Cystinuria/ cystinelysinuria → Cystinosis Cystine storage disorder Cystine accumulates in tissues Deficiency of cystine reductase Death in first 10 years of life.Important: → 4 amino acids are excreted in urine Cystine Ornithine Arginine Lysine → These have common reabsorptive pathway → These have 2 –NH2 groups at almost same distance between → Among these, cystine is most insoluble, hence may form calculi. AROMATIC AMINO ACIDS PHENYL ALANINE AND TYROSINE → Phe Ala essential, tyr non essential → Both glucogenic and ketogenic → Functions of phenyl alanine: Proteins and peptides Converted to tyrosine which has further actions.
O2 H 2OPhenyl alanine tyrosine phe ala hydroxylase activity I tetrahydrobiopterin dihydrobiopterin phe ala hydroxylase activity II (dihydrobiopterin reductase) NADP+ NADPH + H+ Alternate pathway:phe ala phe pyruvate (has keto gp.) phe lactate phe acetate DETOXIFICATION glutamine phe acetyl glutamine → Disorder: phenyl ketonuria Phenyl alanine hydroxylase deficiency During this, above mentioned alternate pathway takes place leading to ketone bodies in urine 1 in 10000 If proper screening is done it is supposed to be 1 in 1500 Diagnosis: • Serum phe ala level: normal <1 mg%, in this disorder, >20 mg% • FeCl3 test FeCl3 + urine green colour presence of phenyl pyruvate
This test is positive in other cases also, hence is only a screening test. → Tyrosine (para hydroxy phe ala) metabolism: transaminaseTyrosine parahydroxy phenyl pyruvate PLP O2 hydroxylase αKG glutamine CO2 Vit C homogentisic acidhomogentisate oxidase O2maleyl acetoacetate isomerase H 2Ofumaryl acetoacetate fumarate + acetoacetate hydrolase glucogenic ketogenicFunctions: → Component of proteins and peptides → Melanin synthesis (from melanocytes)Tyrosine DOPA dopaquinone Tyrosinase, Cu2+ tyrosinase, Cu2+ melanin Quinones of indole hallochrome polymerization spontaneous → Biosynthesis of catecholamines – dopamine, norepinephrine, epinephrine
Tyrosine hydroxylaseTyrosine Dihydroxy Phenyl Alanine (DOPA) Tetrehydrobiopterin dihydrobiopterin PLP DOPA decarboxylase NADP+ NADPH +H+ CO2 DopamineDopamine β hydroxylase, Cu2+ O2Nor epinephrine SAM methyl transferase SAHEpinephrine metanephrine O-methylation Vanillyl Mandelic Acid (VMA) (3- methoxy 4-hydroxy mandelic acid) • Tumours of adrenal medulla, phaeochromocytoma produces high catecholamine levels leading to increased VMA production. → Synthesis of thyroid hormones T3 and T4 Synthesized in follicular cells of thyroid Thyroglobulin has 5000 amino acids, out of which 115 are tyrosine and 35 can be iodinated.Inborn errors of tyrosine metabolism: → Alkaptonuria Deficiency of homogentisate oxidase 1 in 25000
Alkapton is formed from homogentisate, that deposits on connective tissue resulting in ochronosis. Later, may suffer from arthritis NO mental retardation Diagnostic tests: • Urine allowed to stand in urine tube – blackening of urine from above downwards due to oxidation of homogentisic acid. • Positive Benedict’s test • FeCl3 test – green/blue colour.→ Albinism Deficiency of tyrosinase 1 in 20000 Prone to skin cancers.→ Tyrosinemia type I (tyrosinosis) Hepatorenal tyrosinemia Deficiency of fumaryl acetoacetate hydrolase Treatment – diet poor in phenyl alanine and tyrosine→ Tyrosinemia type II (Richner Hanhart syndrome) Occulocutaneous tyrosinemia Deficiency of tyrosine transaminase Formation of palmar keratosis, corneal lesions.→ Neonatal tyrosinemia Deficiency of parahydroxy phenyl pyruvate hydroxylase TRYPTOPHAN→ Indolyl alanine (indole nucleus = benzene +pyrrole rings)→ Essential amino acid→ Both glucogenic and ketogenic→ Products formed: Serotonin Melatonin NAD+
Tryptophan metabolism:It has 11 carbon structure. Out of these 11 C, 1C – formyl group – 1C pool 3C – alanine – glucogenic 4C – acetoacetate – ketogenic 3C – as 3 CO2 O2Tryptophan N-formyl kynurenine Tryptophan pyrrolase THFA Formyl THFA 1C pool Kynurenine when PLP is defecient3 hydroxy kynurenine xanthurenic acid Kynureninase H 2O PLP (VitB6) alanine glucogenic3 hydroxy anthranilic acid NICOTINIC ACID PATHWAY (3%) (97%) Quinolinate aminocarboxy muconaldehyde CO2Nicotinic acid (niacin) amino muconate aldehyde NH3Nicotinate mononucleotide (NMN) ketoadipate CO2Desamido NAD NAD+ acetyl CoA ketogenic
→ Formation of serotonin: TryptophanNADP+ tetrehydrobiopterin tryptophan hydroxylase O2NADPH +H+ dihydrobiopterin H 2O 5 hydroxy tryptophan PLP aromatic amino acid CO2 decarboxylase 5 hydroxy tryptamine (5 HT/ serotonin) Serotonin is excreted as 5 hydroxy indole acetic acid BRANCHED CHAIN AMINO ACIDS VALINE, LEUCINE AND ISOLEUCINEMetabolism:Valine leucine isoleucine PLP transaminaseαketo valenic acid αketo isocaproic acid αketo βmethyl valeric acid branched chain α keto acid dehydrogenaseisobutyryl CoA isovaleryl CoA α methyl butyryl CoA → Maple Syrup Urine Disease(MSUD)/branched chain ketonuria 1 in 100000 Convulsions
Mental retardation Coma, death Onset at 1 month; death in 1 year. valine leucine isoleucine POLYAMINES → Putrescine → Spermidine → SpermineBiosynthesis:Ornithine putrescine spermidine spermine SAM as propylamino group donor and not as methyl donor.Functions: → Production of initiation factors for translation → Cell proliferation → Stabilization of ribosomes and DNA → Synthesis of DNA and RNA → Growth factors, particularly in cell culture systems.Clinical significance: → Increased in cancer tissues → Excretion in urine is increased in cancer.
BIOGENIC AMINES → Produced by decarboxylation of amino acids or their products. → Decarboxylases and PLP are needed. 1. Histidine histamine 2. Ornithine putrescine 3. 5 hydroxy tryptophan 5 hydroxy tryptamine/serotonin 4. DOPA dopamine In intestines by bacteria: 5. Tyrosine tyramine 6. Glycine GABA GLUTAMIC ACID → Acidic → Glucogenic → Non-essentialBiosynthesis: 1. Any amino acid α keto glutarate α keto acid glutamate 2. Histidine, arginine, proline catabolism glutamateCatabolism: NAD+ NADH + H+ 1. Glutamate α keto glutarate L-glutamate dehydrogenase (OXIDATIVE DEAMINATION) 2. By transamination reactions.
Functions: → Component of proteins – mainly gives negative charge → Synthesis of glutathione → Synthesis of GABA → Transport of ammonia → γ carboxy glutamate synthesis in blood clotting factors II, VII, IX, X post-translational modifications Vit K needed → Osteocalcin → N acetyl glutamate GLUTAMINE → Amide of glutamate → Non-essential → GlucogenicBiosynthesis:Glutamate + NH3 glutamineCatabolism:Glutamine glutamate + NH3 GlutaminaseFunctions: → Part of proteins and peptides → Transport of ammonia → Acid base balance → Synthesis of purines and pyrimidines → Conjugating agent.