Metabolism of porphyrins I- Biosynthesis of porphyrins and ... fileRegulation of hemoglobin...

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Metabolism of porphyrins I- Biosynthesis of porphyrins and heme: Hemoglobin is synthesized in erythrocyte precursor cells in bonemarrow as well as in spleen and in liver, The synthesis of protoporphyrin IX occurs in the following steps: i. Formation of δ-aminolevulonic acid (ALA): The starting components for porphyrins biosynthesis are glycine and succinyl CoA that is derived from the citric acid cycle in the mitochondria. Pyridoxal phosphate is also necessary in this reaction to activate glycine. This step is the rate-controlling and is catalyzed by ALA synthase enzyme that occurs in the mitochondria: ii. Formation of porphobilinogen (PBG): A cytoplasmic enzyme, porphobilinogen synthase (ALA-dehydratase), forms porphobilinogen from 2 molecules of ALA. This enzyme is zinc-containing and is sensitive to inhibition by lead:

Transcript of Metabolism of porphyrins I- Biosynthesis of porphyrins and ... fileRegulation of hemoglobin...

Page 1: Metabolism of porphyrins I- Biosynthesis of porphyrins and ... fileRegulation of hemoglobin synthesis: The regulation of heme synthesis occurs at the first step in the pathway, i.e,

Metabolism of porphyrins I- Biosynthesis of porphyrins and heme:

Hemoglobin is synthesized in erythrocyte precursor cells in bonemarrow as well as

in spleen and in liver, The synthesis of protoporphyrin IX occurs in the following

steps:

i. Formation of δ-aminolevulonic acid (ALA):

The starting components for porphyrins biosynthesis are glycine and succinyl CoA

that is derived from the citric acid cycle in the mitochondria. Pyridoxal phosphate is

also necessary in this reaction to activate glycine. This step is the rate-controlling

and is catalyzed by ALA synthase enzyme that occurs in the mitochondria:

ii. Formation of porphobilinogen (PBG):

A cytoplasmic enzyme, porphobilinogen synthase (ALA-dehydratase), forms

porphobilinogen from 2 molecules of ALA. This enzyme is zinc-containing and is

sensitive to inhibition by lead:

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iii. Tetrapyrrole synthesis (Uroporphyrinogen III):

The condensation of 4 PBG units to form uroporphyrinogen III

requires two proteins: a- The enzyme uroporphyrinogen I synthase on its own catalyzes the head-to-

tail condensation of PBG molecules to form uroporphyrinogen I which is of

no physiological importance as it cannot be transferred to heme.

b- A noncatatalytic protein, uroporphyrinogen III cosynthase, alters the

specificity of uroporphyrinogen I synthase, so that one of the PBG

molecules is condensed head-to-head to give uroporphyrinogen III:

iv. Conversion of uroporphyrinogen III to coproporphyrinogen III:

Uroporphyrinogen III is decarboxylated at its aceto side chains by a cytosolic

enzyme to form coproporphyrinogen III with methyl side chain:

v. Conversion of coproporphyrinogen III to protoporphyrinogen III:

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Two mitochondrial enzymes oxidatively decarboxylate 2 side chains to form

protoporphyrinogen III:

vi. Formation of protoporphyrin IX and heme:

Two methylene bridges are oxidized and the formed protoporphyrin IX is

connected with Fe2+ by a mitochonrial enzyme to form heme (protoheme IX):

N.B. Coproporphyrinogen oxidase is only specific to type III coproporphyrinogen, and this

explain why protoporphyrin I has not been identified in natural materials.

Regulation of hemoglobin synthesis:

The regulation of heme synthesis occurs at the first step in the pathway, i.e, at the step

catalyzed by ALA synthase enzyme. This enzyme is allosterically inhibited by protohemin

IX (hemin), which is also corepressor of transcription of ALA synthase gene. Protohemin

IX (hemin) arises from protoheme IX (heme) by oxidation of Fe2+ to Fe3+. This occurs

when heme is in excess of globin because globin-bound heme is stabilized in the Fe2+ state,

but free heme is not. Hemin also regulates the synthesis of globin chains in erythropoietic

cells at the level of mRNA translation.

Abnormal porphyrin biosynthesis (Porphyrias):

Accumulation of porphyrins may occur under conditions of genetically defective syn-

thesis in the liver (hepatic porphyrias) or in the erythropoietic tissues (erythropoietic

porphyrias), as these systems are under seperate genetic control.

1- Acute intermittent hepatic porphyria:

a) It is due to deficiency of uroporphyrinogen I synthase.

Protoporphyrin IX + Fe2 HEME Heme synthase or Ferrocheltase

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b) Excessive amounts of porphobilinogen and ALA are formed.

c) The symptoms are acute abdominal pain and neurologic symptoms.

2- Congenital erythropoietic porphyria:

a) It is due to a decrease of uroporphyrinogen III cosynthase to about one-tenth to one-

third of the normal level.

b) It is characterized by the urinary excretion of large amounts of uroporphrinogen I

c) It causes mutilating skin lesions and hemolytic anemia.

2- Catabolism of heme and formation of bile pigments:

In human, under physiological conditions, about 2x108 erythrocytes are destroyed per

hour. Thus in one day, a 70 kg human turns over about 6 g of hemoglobin. Red cells

destruction occurs usually in the spleen, which is the major site of heme catabolism,

although some occurs in the liver. The catabolism occurs in the microsomal fractions of

the reticuloendothelial cells of spleen or liver by a complex system called heme oxygenase.

When heme reaches the oxygenase system, the iron has usually oxidized to ferric

form (hemin) that may be loosely bound to albumin as methemin. The hemin is reduced

with NADPH, and with the aid of more NADPH an oxygen is added to the α-methyl bridge

between pyrroles I & II. Oxidative opening occurs between rings I & II releasing CO gas

and forms biliverdin which is then reduced to the orange pigment bilirubin as follows:

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Excretion of bilirubin:

1. The resulted bilirubin from heme catabolism is transfered from spleen to liver.

Bilirubin is only sparingly soluble in water, so in plasma it is bound to albumin.

Each mole of albumin appears to have one high-affinity and other low-affinity sites

for bilirubin. In 100 ml of plasma about 25 mg of bilirubin can be tightly bound to

albumin at its high-affinity site. Bilirubin in excess of this quantity can bound only

loosely and thus can be easily detached and diffused into tissues. A number of

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compounds as some antibiotics competes with bilirubin for the high-affinity sites

on albumin and thus can displace bilirubin from albumin.

2. In the liver cell, it is transported through the cytoplasm to the endoplasmicreticulum

bound to a protein called ligand. Here it is conjugated to glucouronic acid by a

specific glucouronyl transferase that transfers the glucuronyl moiety from uridine

diphosphate glucuronate (UDP-glucuronate) to one of the propionyl side chains of

bilirubin. At the plasma membrane, the diglucuronide of bilirubin is formed by the

reaction of 2 moles of bilirubin monoglucuronide to form bilirubin diglucuronide

and free bilirubin. This last reaction is catalyzed by bilirubin glucuroniside

glucuronyl transferase. Then the water soluble bilirubin diglucuronide is secreted

in the bile.

N.B. Conjugated bilirubin (bilirubin diglucuronide) is usually called "direct bilirubin"

(DB) as being assayed directly by diazotization with sulphanilic acid (van den Berg

reaction) in aqueos solution. Free unconjugated bilirubin is called "indirect bilirubin"

(IB) as being diazotized only after methanol addition.

Under normal physiological conditions, most of (DB) in the liver is removed by bile

via the bile duct and excreted to small intestine. As (DB) reaches the terminal ileum and

the large intestine, the glucuronides are removed by specific bacterial enzymes (β-

glucuronidase). It is then reduced by the fecal bacterial flora to a group of colourless

tetrapyrrole compounds called urobilinogens which on air oxidation are oxidized to the

brown pigment urobilin. Some of urobilinogen in the intestine is absorbed to blood and

reexcreted via the liver (enterohepatic circulation) or excreted via kidney (urinary

urobilinogen).

UDP-Glucose UDP-Glucuronic acid

Dehydrogenase

NAD NADHH+

UDP-Glucuronic acid + BILIRUBIN

(IB)

BILIRUBIN monoglucuronide

+ UDP

UDP-Glucuronyl transferase

(endoplasmic reticulum)

2 BILIRUBIN

monoglucuronide BILIRUBIN diglucuronide (DB)

+ BILIRUBIN

Bilirubin glucuroniside

glucuronyl transferase

(plasma membrane)

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When bilirubin in the blood exceeds 1 mg/dl (17.1 µmol/L), hyperbilirubinemia

exists. Hyperbilirubinemia may be due to the production of more bilirubin than the normal

liver can excrete, or it may result from the fai are of a damaged liver to excrete bilirubin

produced in normal amounts. In the absence of hepatic damage, obstruction to the excretory

ducts of the liver, by preventing the excretion of bilirubin, will also cause hyperbilirubi-

nemia. In all these situations, bilirubin accumulates in the blood, and when it reaches a

certain concentration (about 2-2.5 mg/dl), it diffuses into the tissues, which then become

yellow. The condition is called jaundice or icterus.

1- Hemolytic jaundice (prehepatic):

It is due to increased hemolysis of RBC's in a rate faster than normal liver can excrete.

This leads to accumulation of (IB) in serum which usually is abscent in urine being

insoluble. The increased bilirubin leads to increased urobilinogen which appears in large

amounts in urine.

2- Hepatic jaundice:

This type is caused by liver disfunction resulting from damage of parenchymal liver

cells as caused by some hepatotoxic agents e.g carbon tetrachloride and chloroform or

in case of hepatic viral infections. Here both serum (IB) and (DB) are increased. DB

being soluble appears in urine with urobilinogen.

3- Posthepatic or obstructive jaundice:

It is a conjugated hyperbilirubinemia (increased serum DB) results from the blockage

hepatic or bile ducts. (DB) formed in the liver normaly, canmnot be excreted as bile so

absorbed into hepatic vein appears in the circulation. In this case urobilinogen is not

formed and so it is abscent in feces and urine.

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The following table summarizes the laboratory tests that are used to distinguish between

the different types of jaundice, prehepatic, hepatic and posthepatic:

Type Prehepatic Hepatic Poshepatic

Etiology

- Increased hemolysis

of RBC' s at a rate

faster than its

excretion by liver.

- Infective

erythropioesis

- Hepatitis e.g viral or

drug induced.

- Drugs e.g rifampicin

which interfere with

bilirubin conjugation.

- Intrahepatic

obstruction as in

cirrhosis.

- Gilbert's syndrome.

- Poisons as Ccl4, Pb,

As.

- Biliary obstruction

- Gallstone.

- Stricture.

- Carcinoma of

pancreas or

biliary tree.

Total bilirubin Increased Increased Increased

Unconjugated

bilirubin Increased Increased Normal

Conjugated

bilirubin Normal Increased Increased

Urinary

bilimbin Absent Present Present

Urinary

urobilinogen Increased Decreased Absent

Colour of

urine Dark yellow Cola colour Cola colour

Fecal

stercobilinoge

n

Increased Decreased Absent

Colour of stool Dark yellow Light brown Clay colour

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Jaundice in the Newborn:

• Red-cell destruction, together with immature hepatic uptake, conjugation (due to

decreased activity of hepatic glucouronyl transferase) and secretion of bilirubin, may

cause a high plasma level of unconjugated bilirubin in the newborn; so-called

“physiological jaundice” is common.

• As a result of hemolytic disease, the plasma concentration of unconjugated bilirubin

may be as high as 30 mg/dl and exceed the plasma protein-binding capacity; free

unconjugated bilirubin may be deposited in the brain, causing kernicterus, a potential

very serious disorder of the central nervous system.

Treatment:

1. Phototherapy activates hepatic excretion of unconjugated bilirubin which may be due

to converting bilirubin to another geometric isomers that can be more easily excreted

in urine.

2. Administration of Phenobarbital that induces bilirubin-conjugating enzymes.

Some Inherited Hyperbilirubinaemias:

There is a group of inherited disorders in which either unconjugated or conjugated

hyperbilirubinemia is the only detectable abnormality.

1. Gilbert's syndrome is a relatively common, familial condition which may be

present at any age, but usually after the second decade. Plasma unconjugated

bilirubin concentrations are usually between 1.2 and 2.5 mg/dl and rarely exceed 5

mg/dl. They fluctuate and may rise during intercurrent illness or fasting. The

condition is harmless but must be differentiated from haemolysis and hepatitis. It

often becomes evident when plasma bilirubin concentrations fail to return to normal

after an attack of hepatitis, or during any mild illness which, because of the jaundice,

may be misdiagnosed as hepatitis.

2. Crigler-Najjar syndrome, due to deficiency of hepatic glucuronyl transferase, is

more serious. It usually presents at birth. The plasma unconjugated bilirubin

concentration may increase to concentrations that exceed the binding capacity of

albumin and so cause kernicterus.