Keshava Pavan K, medical student, KMC, Mangalore, India

AMINO ACIDS AND PROTEINS

Important 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:

→ Transamination

Alanine α ketoglutarate

PLP ALT

Pyruvate glutamate

PLP – pyridoxal phosphate; ALT – alanine transaminase/SGPT -serum glutamate

pyruvate transaminase

→ Trans deamination (transamination + deamination)

Aspartate α ketoglutarate

PLP AST

Oxaloacetate glutamate

NAD+/NADP+

Oxidative demination in liver glutamate dehydrogenase

NADH+ H+/NADPH+ H+

α ketoglutarate + NH4+

AST – aspartate transaminase/SGOT serum glutamate oxaloacetate transaminase

→ Oxidative deamination

L amino acid α keto acid + NH4+

FMN FMNH2

D amino acid α keto acid + NH4+

FAD FADH2

→ Non oxidative deamination

Serine pyruvate + NH4+

Threonine α keto butarate + NH4+

oxidase

oxidase

Serine dehydratase

Threonine dehydratase

Cysteine pyruvate + H2S + NH4+

Glutamate + NH4+ glutamine

In liver glutaminase

ATP ADP + Pi glutamate + NH4+

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 DEFECIENCY

Hyperammonemia type I CPS I

Hyperammonemia type II OTC

Citrullinemia ASS

Argininosuccinic aciduria ASL

Hyperargininemia Arginase

→ Normal serum urea level: 20 to 40 mg/dL

→ Increase in serum urea level – uremia

→ Causes:

Cysteine desulphydratase

Glutamine synthetase

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

Histidine histamine + CO2

5-hydroxy tryptophan 5-hydroxy tryptamine/ serotonin

Glutamate γ amino butyric acid

SIMPLEST AMINO ACID

GLYCINE

→ Glucogenic

→ Non essential

→ Synthesis :

PLP

Histidine decarboxylase

FH4 N5 N10 methylene FH4

Serine glycine

Serine hydroxy

methyl transferase

Threonine glycine + acetaldehyde

Threonine aldolase

→ Catabolism:

NAD+ NADH

PLP

Glycine CO2 + NH4+

FH4 N5N10 methylene FH4

The reverse reaction is used in synthesis of glycine. Enzyme is then named glycine

synthase.

Deficiency of enzymes of glycine cleavage system causes non-ketotic hyperglycemia

Glycine serine pyruvate gluconeogenesis

Glycine

cleavage

system

Serine dehydratase

Malate

Glycine glyoxalate Oxalate

Glycine oxidase Formate

Block 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 +Pi

Glutamate + 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

NH4+

Cholic acid + glycine glycocholic acid

Chenodeoxy cholic acid + glycine glyco chenodeoxycholic acid

Benzoic 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 SAH

Guanidoacetate creatine

Guanidoacetate methyl transferase

Norepinephrine epinephrine

Nicotinamide 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 synthase

Homocysteine + serine cystathionine

PLP H2O cystathionase

H2O

PLP

Homoserine + cysteine

→ Glucogenic pathway

Homoserine α keto butyrate propionic acid

NH2 CO2

Glucogenic TCA cycle succinyl CoA

Methionine α keto γ methiol butyrate

α keto butyrate + methyl mercaptan (CH3 – SH)

→ Inborn errors of methionine metabolism:

→ Homocystinuria

2 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

2H

2 cysteine cystine

Cysteine reductase 2H

→ Biosynthesis: refer trans-sulphuration pathway under methionine.

→ Catabolism:

Transamination

PLP

Cysteine mercaptopyruvate H2S + pyruvate glucogenic

α KG glu

cysteine desulphydratase

cysteine + H2O pyruvate + NH3 + H2S

cysteine dioxygenase

cysteine cysteine sulfinic acid

2O2 2H2O α KG

2NADH 2NAD+ PLP

Glu

Sulfinyl pyruvate

Desulfinase

Pyruvate + sulphurous acid

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.

→ Detoxification

Bromobenzene 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 H2O

Phenyl 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:

transaminase

Tyrosine parahydroxy phenyl pyruvate

PLP O2 hydroxylase

αKG glutamine CO2 Vit C

homogentisic acid

homogentisate oxidase O2

maleyl acetoacetate

isomerase H2O

fumaryl acetoacetate fumarate + acetoacetate

hydrolase

glucogenic ketogenic

Functions:

→ 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 hydroxylase

Tyrosine Dihydroxy Phenyl Alanine (DOPA)

Tetrehydrobiopterin dihydrobiopterin PLP

DOPA decarboxylase

NADP+ NADPH +H+ CO2

Dopamine

Dopamine β hydroxylase, Cu2+ O2

Nor epinephrine

SAM

methyl transferase

SAH

Epinephrine 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

O2

Tryptophan N-formyl kynurenine

Tryptophan pyrrolase THFA

Formyl THFA 1C pool

Kynurenine

when PLP is defecient

3 hydroxy kynurenine xanthurenic acid

Kynureninase H2O

PLP (VitB6) alanine glucogenic

3 hydroxy anthranilic acid

NICOTINIC ACID PATHWAY (3%) (97%)

Quinolinate aminocarboxy muconaldehyde

CO2

Nicotinic acid (niacin) amino muconate aldehyde

NH3

Nicotinate mononucleotide (NMN) ketoadipate

CO2

Desamido NAD NAD+ acetyl CoA ketogenic

→ Formation of serotonin:

Tryptophan

NADP+ tetrehydrobiopterin tryptophan hydroxylase

O2

NADPH +H+ dihydrobiopterin H2O

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 ISOLEUCINE

Metabolism:

Valine leucine isoleucine

PLP transaminase

αketo valenic acid αketo isocaproic acid αketo βmethyl valeric acid

branched chain α keto acid dehydrogenase

isobutyryl 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

→ Spermine

Biosynthesis:

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-essential

Biosynthesis:

1. Any amino acid α keto glutarate

α keto acid glutamate

2. Histidine, arginine, proline catabolism glutamate

Catabolism:

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

→ Glucogenic

Biosynthesis:

Glutamate + NH3 glutamine

Catabolism:

Glutamine glutamate + NH3

Glutaminase

Functions:

→ Part of proteins and peptides

→ Transport of ammonia

→ Acid base balance

→ Synthesis of purines and pyrimidines

→ Conjugating agent.

Diagram (1)

Diagram (2)

Diagram (3)

Diagram (4)

Diagram (5)

Diagram (6)

Diagram (7)

Diagram (8)

Diagram (9)

Diagram (10)

Diagram (11)