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Biosynthesis of Heme
Dr.S.Chakravarty ,MD
Biosynthesis of Heme
• TISSUES :- ALL TISSUES BUT PREDOMINANTLY IN THE LIVER AND BONE-MARROW
• SUBCELLULAR SITE :- PARTLY IN MITOCHONDRION ( FIRST STEP AND THE LAST THREE STEPS )
AND
PARTLY CYTOSOLIC.(REMAINING STEPS)
• STARTING MATERIALS :- 1)Succinyl CoA( FROM TCA CYCLE) 2)AA - GLYCINE
MITOCHONDRIA
Lead Inhibits
1
2
MITOCHONDRIA
COMMITTED STEP & RATE –LIMITING
3
HYDROXYMETHYLBILANE(LINEAR TETRAPYRROLE)
FOUR MOLECUES OF PRORPHOBILINOGEN
TYPE III UROPORPHYRINOGENTYPE I UROPORPHYRINOGEN
4
Also called PBG deaminase
5
ACETYL TO METHYL ( AM )
Coporporphyrinogen oxidase
Protoporphyrinogenoxidase
Ferrochelatase
COPROPORPHYRINOGEN III
PROTOPORPHYRINOGEN III
PROTOPORPHYRIN III
Fe +2
HEME
M
I T
O C
H
O N
D R
I A
6
7
8
PROPIONYL TO VINYL(PV ) 2CO2
INCORPORATION OF IRON TO PROTOPORPHYRIN
Iron is coordinately linked with 5N (4 Pyrrole and 1 N2 of a His residue of Globin)
Lead inhibits
Uroporphyrinogen I synthase
Uroporphyrinogen III synthase
Uroporphyrinogen decarboxylase
Coporporphyrinogen oxidase
Protoporphyrinogenoxidase
Ferrochelatase
6
7
8MIT
OCH
ON
DRI
ACY
TOSO
L
• REMEMBER :-
– 2/3 rd of TOTAL HEME SYNTHESIZED GOES FOR SYNTHESIS OF cytP450.
–cytP450 is involved in metabolism of drugs and xenobiotics.
Regulation
• The rate limiting enzyme is ALA SYNTHASE which exists in two forms ALAS1(HEPATIC ) & ALAS2 (ERYTHROID )
Feedback Regulation of ALAS1:-
• HEME with the help of a APO-REPRESSOR acts as a NEGATIVE REGULATOR.
• So, the Rate of synthesis of ALA SYNTHASE greatly increases in the absence of heme and is diminished in its presence.
FEEDBACK
INHIBITION
• MANY DRUGS CAN CAUSE MARKEDLY INCREASE ALA SYNTHASE . -WHY ?
• Several factors can cause DEREPRESSION OF ALAS-1 IN LIVER eg. – Glucose administration – Administration of HEMATIN .
• ALAS-2 DOESNOT UNDERGO FEEDBACK REGULATION BY HEME .
• ALAS-2 IS NOT INDUCED BY DRUGS THAT INDUCE ALAS-1.
Porphyrins are coloured and Fluoresce
• The Porphyrinogens are colourless, whereas the various Porphyrins and are coloured.
• There is a characteristic absorption curve for each porphyrin. For eg.( in 5%HCl )
Peak absorption of hematoporphyrins at 400nm
SORET BAND
The Porphyrias• The porphyrias are a group of genetic disorders due to
abnormalities in the pathway of biosynthesis of heme.
• The symptoms produced depend on the type of defect.
• Autosomal dominant porphyrias include acute intermittent porphyria, most cases of erythropoietic protoporphyria, hereditary coproporphyria, and variegate porphyria.
• Most common is PORPHYRIA is AIP
• If the lesion occurs early in the pathway prior to the formation of porphyrinogens ALA and PBG will accumulate in body tissues and fluids .– Clinically they will complain of abdominal pain and
neuropsychiatric symptoms.
• If the enzyme is blocked later in the pathway it will lead to accumulation of porphyrinogens .
Porphyrinogens
2H
Porphyrin derivatives
PHOTOSENSITIVITY
– The porphyrins when exposed to light of 400nm become excited and then react with molecular oxygen to produce oxygen radicals.
– These radicals injure lysosomes and other cellular organelle.
The lysosomes release degradative enzymes which cause various damage to skin , resulting in SCARRING and damage to organs like the Liver.
Spontaneous oxidation
Light
Summary of Major Findings in the Porphyrias
Enzyme Involved Type and Class Major Signs and Symptoms
Results of Laboratory Tests
1. ALA synthase (erythroid form)
X-linked sideroblastic
anemia(erythropoietic)
Anemia ONLY . Red cell counts and hemoglobin
decreased
2. ALA dehydratase
ALA dehydratase deficiency (hepatic)
Abdominal pain, neuropsychiatric
symptoms(A-N )
Urinary ALA
3. Uroporphyrinogen I synthase
Acute intermittent
porphyria (hepatic)
Abdominal pain, neuropsychiatric
symptoms(A-N)
Urinary ALA and PBG increased
Enzyme Involved Type and Class Major Signs and Symptoms
Results of Laboratory Tests
4.Uroporphyrinogen III synthase
Congenital erythropoietic
(erythropoietic)
Photosensitivity(P)
ALA , PBG ++
5. Uroporphyrinogen decarboxylase
Porphyria cutanea tarda (hepatic)
Photosensitivity(P)
Urinary uroporphyrin I increased
6. Coproporphyrinogen oxidase
Hereditary coproporphyria
(hepatic)
Photosensitivity, abdominal pain, neuropsychiatric
symptoms(P-A-N)
Urinary ALA, PBG, and coproporphyrin
III and fecal coproporphyrin III
increased7. Protoporphyrinogen oxidase
Variegate porphyria (hepatic)
Photosensitivity, abdominal pain, neuropsychiatric
symptoms(P-A-N)
Urinary ALA, PBG, and coproporphyrin
III and fecal protoporphyrin IX
increased8. Ferrochelatase Protoporphyria
(erythropoietic)Photosensitivity
(P)Fecal and red cell protoporphyrin IX
increased
• Acute porphyrias• Central Nervous system,
– resulting in abdominal pain, vomiting, acute neuropathy, muscle weakness, seizures
– mental disturbances, including hallucinations, depression, anxiety, and paranoia.
• SEVERE PAIN – BOTH ACUTE AND CHRONIC
• Constipation is frequently present, as the nervous system of the gut is affected, but diarrhea can also occur.
• Patients with acute porphyria (AIP, HCP, VP) are at increased risk over their life for hepatocellular carcinoma (primary liver cancer) and may require monitoring. Other typical risk factors for liver cancer need not be present.
Cutaneous pophyrias
– Photosensitivity and skin lesions
Disorder Enzyme defect
Neurovisceral crises
Skin lesions
Inheritance OMIM#
Acute porphyriasALA dehydratase deficiencyporphyria (ADP)
ALAD + - AR 125270
Acute intermittent porphyria (AIP)
PBGD + - AD 176000
Hereditary coproporphyria (HCP)
CPO + +1,2 AD 121300
Variegate porphyria (VP) PPOX + +1,2 AD 600923Non-acute porphyriasCongenital erythropoietic porphyria (CEP)
UROS - +2 AR 263700
Porphyria cutanea tarda (PCT)
UROD - +2 Complex 176090, 176100
Erythropoietic protoporphyria (EPP)
FECH - +3 AD 177000
1Skin lesions and neurovisceral crises may occur alone or together; 2fragile skin, blisters; 3acute photosensitivity without fragile skin, blisters.
PORPHYRIA CUTANEA TARDA
GUNTHER’S DISEASE(Congenital Erythropoietic porphyria )
GUNTHER’S DISEASE(Congenital Erythropoietic porphyria )
Gingival recession
GUNTHER’S DISEASE(Congenital Erythropoietic porphyria )
Erythrodontia
Why accumulation PBG and ALA lead to complications:
• ALA and PBG are Neurotoxic
• Vasoconstriction of CNS and abdominal blood vessels.
• Decreased Heme containing proteins - Decreased cellular energy.
• Elevation of stress hormones – Anxiety, Increase in BP etc.
PRECIPITATING FEATURES• DRUGS :- Certain medicines, such as
BARBITURATES(SEDATIVE ) , GRISEOFULVIN (ANTI-FUNGAL)birth control pills, antibiotics, and medicines for treating seizures.
• Why ? THEY ARE METABOLISED BY CYT P450 , hence UTILIZATION of HEME in CYTOCHROME P450 IS INCREASED .This DECREASES THE INTRACELLULAR CONCENTRATION.
• Dehydration (loss of too much water and salt) too much sun exposure.
• Cigarette smoking and alcohol consumption.
• Hormone changes,during menses , menarche and menopause.
• Physical or mental stress, such as with an infection, depression, emotional problem, or after surgery.
• Starvation due to fasting or crash dieting.
Why accumulation PBG and ALA lead to complications:
• ALA and PBG are Neurotoxic
• Vasoconstriction of CNS and abdominal blood vessels.
• Decreased Heme containing proteins - Decreased cellular energy.
• Elevation of stress hormones – Anxiety, Increase in BP etc.
DIAGNOSIS• Porphyria is diagnosed through spectroscopy and
biochemical analysis of blood, urine, and stool.
• In general, urine estimation of porphobilinogen (PBG) is the first step if acute porphyria is suspected.
• In nearly all cases of acute porphyria syndromes, urinary PBG is markedly elevated except for the very rare ALA dehydratase deficiency.
• Urinary ALA and PBG are always markedly increased in symptomatic patients with AIP and even in some asymptomatic individuals with the inherited enzyme deficiency.
Diagnosis
PBG in urine is oxidized to porphobilin upon standing, which gives a dark-brown color to urine, and often referred to as ‘port-wine reddish urine’.
Lab diagnosis:
• Urinary PBG excretion during an attack :– usually 50–200 mg/24 h [normal, 0–4mg/24 h]
• Urinary ALA excretion : – 20–100 mg/24 h
[normal, 1–7 mg/24 h]
• Portwine color of urine on prolonged standing
• Repeat testing during an attack and subsequent attacks may be necessary in order to detect a porphyria, as levels may be normal or near-normal between attacks.
• UV fluorescence :- Presence of porphyrins in urine is detected by Ehrlich’s reagent. If urine is observed under UV light it emits strong red-fluorescence if porphyrins are present.
• DNA analysis to identify the causative mutation in the appropriate gene (AIP:HMBS; VP: PPOX; HCP: CPO ) is the method of choice.
• Enzyme measurements are not as specific or sensitive as DNA analysis. – Erythrocyte PBG deaminase assay is still used for detection of latent AIP
when DNA analysis is not available or a mutation cannot be detected.– Measurement of protoporphyrinogen and coproporphyringen oxidases is
complex and requires nucleated cells.
Additional Tests
• USG of LIVER AND LIVER ENZYMES :- predisposed to Hepatocellular Carcinoma
• NERVE CONDUCTION STUDIES :- PERIPHERAL NEUROPATHY due to axonal degeneration affects proximal muscles esp. shoulders and arms and may lead to Bulbar paralysis DEATH if affected.
TREATMENT
• Acute phase :- • I.V GLUCOSE- GLUCOSE
REPRESSES ALA SYNTHASE • I.V HEMATIN –HEME REPRESSES
ALA SYNTHASE• PAIN MANAGEMENT :-
Narcotics • NAUSEA AND VOMITING :- PHENOTHIAZINES • SEDATION • MANAGEMENT OF SEIZURES :- Clonazepam
• SUSCEPTIBLE PATIENTS MUST AVOID DIRECT EXPOSURE TO SUNLIGHT –sunscreens may help.
ACQUIRED PORPHYRIAS
Porphyrinuria can occur in Lead Poisoning due to inhibition of ferrochelatase also.
Rem :- Lead also inhibits ALA dehydratase
MCQ• AIP is the major autosomal-dominant acute hepatic porphyria.
This disease is caused by a deficiency in porphobilinogen (PBG) deaminase, an enzyme of heme biosynthesis. Patients afflicted with this disease would be expected to excrete excess amounts of which of the following?
(A) Delta-aminolevulinic acid (ALA)(B) Coproporphyrinogen III(C) Hydroxymethylbilane(D) Protoporphyrin IX(E) Type III uroporphyrinogen
Uroporphyrinogen decarboxylase
ALA DEHYDRATASE
• MCQ 1
• A 25-year-old man has experienced chronic blistering and scarring of his skin when exposed to sunlight. This man is a smoker and drinks heavily, both of which exacerbate his responses to sunlight. Analysis of his urine and plasma indicates a high accumulation of complex porphyrins, predominantly uroporphyrin.The symptoms and clinical signs displayed by this patient indicate he is suffering from which of the following disorders?
(A) Acute intermittent porphyria (AIP)(B) Hereditary coproporphyria (HCP)(C) Porphyria cutanea tarda (PCT)(D) Variegate porphyria(E) X-linked sideroblastic anemia