porphyrin metabolism

Question 1

porphyrins

Answer 1

cyclic compounds that bind metal ions tightly
most important = heme
formed by linkage of 4 pyrrole rings through methenyl bridges
two double bonds are conjugated throughout system
ones in humans have asymmetrical rings - can have symmetrical ones in disorders

Question 2

heme

Answer 2

iron-binding compound responsible for:

- O2 transport in blood and muscle
- electron transport in mitochondria
- metabolism of fat-soluble compounds in the liver

Question 3

pyrrole rings

Answer 3

form porphyrins
each ring has two side chains which are distributed around entire porphyrin molecule
Roman numerals specify the arrangement of side chains - asymmetrical when porphyrin is produced under normal physiological conditions

Question 4

central pocket

Answer 4

of a porphyrin
can be occupied by a metal ion
iron is in heme
cobalt is in cobalamine (vitamin B12)
magnesium is in the chlorophyll in plants

Question 5

physiological roles of heme proteins (list)

Answer 5

as mitochondrial cytochromes - in electron transport chain
in Hb and Mb to transport O2
as catalases
in cytochrome P450 proteins - drug metabolism
removal of H2O2 - catalase

Question 6

heme proteins as mitochondrial cytochromes

what inhibits?

Answer 6

generate membrane H+ gradient required for ATP synthesis via electron transport chain
cyanide poisoning causes irreversible inhibition of mitochondrial cytochrome a3

Question 7

heme in cytochrome P450 enzymes

Answer 7

needed for metabolism of fat-soluble compounds, formation of cholesterol, steroids, and arachidonic acid metabolites

Question 8

heme in catalase

Answer 8

antioxidant enzyme hydrolyzes H2O2

Question 9

steps in heme synthesis

Answer 9

1: ALA synthase catalyzes condensation of glycine and succinyl CoA to make delta-aminolevulinic acid (ALA)
2: ALA dehydrase condenses two molecules of ALA to make pyrrole compound porphobilinogen (PBG)
3: hydroxymethylbilane synthase condenses four molecules of PBG to make hydroxymethylbilane
4: uroporphyrinogen synthase catalyzes ring closure of hydroxymethylbilane to make uroporphyrinogen I
5: uroporphyrinogen III cosynthase isomerizes the D ring side chains of uroporphyrinogen I to make uroporhyrinogen III
6: decarboxylation and oxidation reactions convert uroporphyrinogen III to protoporphyrin IX
7: ferrochelatase catalyzes introduction of iron into protoporphyrin IX to make heme

Question 10

where is heme synthesized?

Answer 10

primarily by liver and bone marrow specific isoforms of the necessary enzymes

Question 11

ALA synthase

Answer 11

enzyme that catalyzes the first step of heme synthesis
condenses glycine and succinyl CoA to make delta-aminolevulininc acid (ALA)
releases CoA and CO2
irreversible reaction
hemin, oxidized form of heme (Fe3+), and glucose suppress transcription of ALA synthase gene - allows negatative feedback

Question 12

what suppresses ALA synthase?

Answer 12

hemin, oxidized form of heme, and glucose

note: these suppress the synthesis of the mRNA, not the activity of the enzyme

Question 13

ALA dehydrase

Answer 13

enzyme responsible for second step of heme synthesis = pyrole compound
condenses 2 molecules of ALA to make porphobilinogen (PBG)
releases two H2O
lead inhibits

Question 14

hydroxymethylbilane synthase

Answer 14

enzyme responsible for 3rd step in heme synthesis
condenses 4 molecules of PBG to make hydroxymethylbilane
regulate reaction
second rate-limiting step
releases 4NH3

Question 15

uroporphyrinogen synthase

Answer 15

enzyme responsible for 4th step in heme synthesis

catalyzes ring closure of hydroxymethylbilane to make uroporphyrinogen I - symmetrical

Question 16

uroporphyrinogen III cosynthase

Answer 16

enzyme responsible for 5th step in heme synthesis
isomerizes the D ring side chains of uroporphyrinogen I to make uroporphyrinogen III - so that it’s not a symmetrical heme and can be used biologically

Question 17

uroporphyrinogen III

Answer 17

molecule made in 5th step of heme synthesis by enzyme uroporphyrinogen III cosynthase from uroporphyrinogen I
common precursor for chlorophyll, cobalamine, and heme in organisms that make those compounds
several steps make it into protoporphyrin IX, which has lots of conjugated double bonds that allow them to fluoresce

Question 18

how could you tell the difference between porphyrins and their precursors?

Answer 18

porphyrins are purple and fluorescent

PBG and other porphyrinogens are colorless and don’t fluoresce until they’re oxidized

Question 19

ferrochelatase

Answer 19

enzyme responsible for 7th step of heme synthesis
catalyzes the introduction of iron (Fe2+) into protoporphyrin IX to make heme
lead inhibits
releases 2 H+

Question 20

classes of disease associated with abnormal heme synthesis (4 - list)

Answer 20

1: acute porphyrias
2: non-acute porphyrias
3: lead poisoning
4: iron-deficiency anemia

Question 21

what steps of heme synthesis are disrupted by lead?

Answer 21

the enzymes ALA dehydrase and ferrochalatase are inhibited by lead
the first condenses two molecules of ALA to make PBG
the second catalyzes the addition of iron to make the final heme

Question 22

acute porphyrias

Answer 22

autosomal dominant disorders characterized by blockade of the early, rate-limiting steps of heme biosynthetic pathway
results in decreased production of heme (and hemin)
example = acute intermittent porphyria (AIP)

Question 23

acute intermittent porphyria (AIP) (everything we’ve learned about it)

Answer 23

due to deficiency in hydroxymethylbilane synthase
clinical signs: intermittent attacks of abdominal pain and neuropsychiatric symptoms (that go away with bowel movements)
attack precipitated by ingestion of drugs and other compounds
compounds induce production of CYP enzymes => consumption of heme => increase in ALA synthase levels and accumulation of ALA and PBG - ALA committed step
lack of negative feed back makes things worse
ALA causes abdominal pain because of its massive accumulation in the liver
neuropsychiatric symptoms due to similarity between ALA and GABA => ALA antagonizes GABA receptors
diagnosis is made by detection of excess PBG in urine - when urine stands, it oxidizes and turns purple
treat with intravenous hemin and glucose
patients should avoid precipitating drugs
can identify affected family members by PCR

Question 24

what deficiency causes acute intermittent porphyria?

what is the result of this deficiency (what will build up)?

Answer 24

deficiency of hydroxymethylbiane synthase enzyme, which condenses 4 molecules of PBG to make hydroxymethylbilane

this will result in the build up of PBG and ALA (the precursor for PBG)

Question 25

what are the clinical symptoms of acute intermittent porphyria (AIP)?

Answer 25

abdominal pain | neuropsyciatric symptoms

Question 26

what usually triggers an attack of acute intermittent porphyria (AIP)? why does this trigger an attack?

Answer 26

attacks are usually triggered by ingestion of drugs and other symptoms this is because these compounds induce the production of CYP enzymes => heme consumption => increases in ALA synthase activity => increases in ALA and PBG the ALA and PBG builds up in liver patients with AIP are advised to avoid precipitating drugs

Question 27

what causes the abdominal pain that patients with acute intermittent porphyria (AIP) experience?

Answer 27

buildup of ALA in the liver

Question 28

what causes the neuropsyciatric symptoms that patients with acute intermittent porphyria (AIP) experience?

Answer 28

ALA and the neurotrasmitter GABA have very similar structures ALA antagonzies GABA receptors

Question 29

how do you diagnose acute intermittent porphyria (AIP)?

Answer 29

detect excess PBG in urine leave urine to stand so that it can oxidize if the patient has AIP, the urine will turn purple (because when the PBG oxidizes it becomes porphyrin, which is purple)

Question 30

how do you treat acute intermittent porphyria (AIP)?

Answer 30

with intravenous hemin and glucose - ala synthase also negatively regulated by glucose (get rid of/avoid inducers - cytochrome P450 can't work - so anything that needs those) can consume blood - will alleviate symptoms

Question 31

how would you identify family members with acute intermittent porphyria (AIP)?

Answer 31

by PCR

Question 32

non-acute porphyrias (summary card - everything we've learned)

Answer 32

primary acquired diseases associated with liver damage some are genetic blockade of normal heme biosynthetic pathway occurs at specific steps beyond the formation of hydroxymethylbbilane results in accumulation of of porphyrin derivatives in liver and skin => liver damage and rashes heme production and ALA levels are normal so no neuropsychiatric symptoms example = porphyria cutanea tarda (PCT)

Question 33

what is the defect that causes non-acute (chronic) porphyrias? why does this occur?

Answer 33

blockade of normal heme biosynthetic pathway at specific steps beyond the formation of hydroxymethylbilane uroporphyrinogen can't be converted to protoporhyrin IX usually not genetic - due to liver damage

Question 34

what accumulates in non-acute porphyrias? | which tissues are affected, and what does the accumulation cause in these tissues?

Answer 34

abnormal uroporphyrinogen turns into uroporphyrins these derivates accumulate in the liver and the skin in the liver it causes liver damage in the skin it causes photosensitive skin rashes urine turns red

Question 35

what are heme and ALA levels in patients with non-acute porphyria?

Answer 35

both levels will be normal because both ALA synthase and hydroxymethylbilane synthase activities are increased through compensatory regulation mechanisms

Question 36

are there neurophysiological symptoms with non-acute porphyrias?

Answer 36

no. since ALA is normal, there will be no neurological effects

Question 37

porphyria cutanea tarda (PCT) (summary card)

Answer 37

most common porphyria form of non-acute porphyria due to reduced uroporphyrinogen decarboxylase activity => accumulation of uroporphyrins treat with regular phlebotomy => helps remove excess porphyrin metabolites patients should avoid alcohol, sunlight, other liver toxins

Question 38

what defect causes porphyria cutanea tarda (PCT)? | what accumulates as a result?

Answer 38

reduced uroporphyrinogen decarboxylase activity results in the buildup of uroporphyrins, which are abnormal metabolites

Question 39

how is porphyria cutanea tarda (PCT) treated?

Answer 39

with regular phlebotomy, which helps remove excess porphyrin metabolites

Question 40

what should patients with porphyria cutanea tarda (PCT) avoid?

Answer 40

alcohol other liver toxins excess sunlight

Question 41

what enzymes are inhibited in lead poisoning and what symptoms does this cause?

Answer 41

inhibits ALA dehydrase and ferrochelatase clinical symptoms include all symptoms of AIP (due to inhibition of liver ALA dehydrase) plus anemia (due to inhibition of bone marrow heme synthesis)

Question 42

iron-deficiency anemia (summary card)

Answer 42

some of the heme-synthesis enzymes are erythroid specific isoforms and so are regulated by different signals than their liver counterparts eg translation of erythroid ALA synthase mRNA is stimulated by iron lack of iron results in decrease of erythroid heme synthesis => anemia ALA synthase mRNA has iron response element (IRE) in transcript

Question 43

how does iron affect translation of enzymes involved in heme synthesis?

Answer 43

translation of some of the bone marrow enzymes is regulated by iron, especially erythroid ALA synthase the transcript for ALA synthase contains and iron response element (IRE) sequence - makes it susceptible to regulation by iron - if iron not present, the mRNA is blocked from being translated

Question 44

how does lack of iron result in iron-deficiency anemia?

Answer 44

since regulation of ALA synthase mRNA is regulated by iron (through the IRE), lack of iron can prevent the protein from being transcribed, preventing synthesis of heme in bone marrow and causing anemia

Question 45

where does heme degradation occur?

Answer 45

primarily in the spleen and the liver

Question 46

steps of heme degradation:

Answer 46

1: heme oxidase catalyzes conversion of heme to biliverdin, using NADPH and O2 2: heme oxidase also catalyzes release of bound iron molecule when heme ring in broken 3: bilirubin reductase converts biliverdin to bilirubin 4: serum albumin carries the bilirubin through the bloodstream to the liver 5: bilirubin glucoronyltransferase (in liver) conjugates 2 molecules of glucuronic acid to bilirubin to make bilirubin diglucuronide 6: bilirubin diglucuronide is water soluble and is secreted in bile 7: bacteria in the intestine reduce bilirubin diglucuronide to make urobilinogen 8a: most of urobilinogen converted to stercobilin - brown compound that give feces color 8b: rest transported back to liver where converted to urobilin - yellow - excreted in urine

Question 47

heme oxygenase

Answer 47

enzyme that catalyzes the first two steps of heme degradation uses NADPH and O2 to convert heme to biliverdin (green pigment) and iron molecule is removed when ring is broken - linearlizes releases carbon monoxide

Question 48

biliverdin reductase

Answer 48

enzyme responsible for 3rd step in heme degradation uses NADPH reduces biliverdin to make bilirubin (fat-soluble, yellow-red pigment) bound to albumin for transport

Question 49

bilirubin glucuronyltransferase

Answer 49

enzyme that catalyzes the 5th step in heme degradation in liver conjugates 2 molecules of glucuronic acid to bilirubin - makes bilirubin diglucuronide (conjugated bilirubin) makes bilirubin water soluble so can now just move through the blood

Question 50

bilirubin diglucuroninde

Answer 50

water soluble compound made during the degradation of heme made in liver and secreted in bile made by bilirubin glucoronyltransferase from glucuronic acid and bilirubin converted to urobilinogen in intestine by bacteria

Question 51

jaundice (summary card)

Answer 51

due to disorders that increase levels of bilirubin in the blood results in yellow skin and sclerae bilirubin can cross BBB - cause encephalopathy measured with van den Bergh reaction 4 types of jaundice: hemolytic obstructive hepatocellular neonatal if can accumulate to high enough level, can cross BBB and cause coma

Question 52

what are the symptoms of jaundice?

Answer 52

bilirubin builds up in the skin and sclerae, turns them yellow can cross BBB and cause encephalopathy

Question 53

what assay is used to measure levels of bilirubin in the blood?

Answer 53

the van den bergh reaction | measures levels of both conjucated and unconjugated bilirubin in the blood

Question 54

types of jaundice (4 - list)

Answer 54

1: hemolytic 2: obstructive 3: hepatocellular 4: neonatal jaundice

Question 55

hemolytic jaundice - cause? - van den bergh assay results will show?

Answer 55

red cell lysis, e.g. in sickle cell anemia | increase in unconjugated bilirubin

Question 56

obstructive jaundice - cause? - van den Bergh test will show?

Answer 56

blockage of the bile ducts, e.g. by gallstones | increase in conjugated bilirubin

Question 57

hepatocellular jaundice - cause? - how to test and what will the results be?

Answer 57

due to liver damage e.g. by hepatitis | use liver tests - will see increase in AST and ALT

Question 58

neonatal jaundice - causes? - treatment?

Answer 58

due to relatively low levels of bilirubin glucuronyltransferase activity early in infancy - this enzyme isn't expressed until birth/full term so if born early won't be expressing the enzyme yet treated by administration of blue fluorescent light to convert bilirubin to water-soluble molecules that can be excreted by the kidney

Question 59

steps of heme synthesis (molecules made)

Answer 59

1: succinyl CoA + glycine 2: ALA 3: PBG 4: hydroxymethylbilane 5: uroporphyrinogen 6: protoporhyrin IX 7: heme

Question 60

how would you diagnose lead poisoning?

Answer 60

basophillic strippling on blood smear

Question 61

how would you diagnose iron deficiency anemia?

Answer 61

blood smear shows pale, thin rimmed RBCs

Question 62

degradation of heme (molecules made and where)

Answer 62

1: heme - RBCs 2: biliverdin - macrophages, esp in spleen 3: bilirubin - macrophages, esp in spleen 4: conjugated bilirubin - liver 5: urobilinogen - intestine 6a: stercobilin - intestine 6b: urobilin - kidney

Question 63

how would you distinguish between producing too much bilirubin to not being able to get it into the intestine?

Answer 63

ask about color of urine and stool liver function tests - for enzymes that only live in liver - look for damage to liver cells van den bergh reaction

Question 64

van den bergh reaction

Answer 64

dissolve bilirubin in water and methanol only conjugated bilirubin is dissolvable in water dissolving in methanol will give total amount of bilirubin can see how much is being conjugated

Question 65

why would we produce too much bilirubin?

Answer 65

high turnover of RBCs: sickle cell anemia, phalycemia causes hemolytic jaundice - high levels of unconjugated bilirubin

Question 66

why would we have too much conjugated bilirubin?

Answer 66

gall stone obstruction jaundice inability of bilirubin to get into intesting pale stools

Question 67

how would you diagnose hepatocellular jaundice?

Answer 67

get rise in conjugated bilirubin and overall bilirubin - use van den bergh reaction measure with liver enzyme tests - high levels of AST and ALT

Question 68

types of jaundice (summary card)

Answer 68

``` 1: hemolytic - eg sickle cell anemia high levels of unconjugated bilirubin 2: obstructive jaundice - eg gallstones high levels of conjugated bilirubin 3: hepatocellular jaundice - eg hepatits high levels of both conjugated and unconjugated bilirubin 4: neonatal jaundice - premature infants high levels of unconjugated bilirubin ```

Question 69

pyridoxal phosphate (PLP)

Answer 69

coenzyme required by ALA synthase to catalyze the first step of porphyrin synthesis

Question 70

sideroblastic anemia (lippincott)

Answer 70

loss of function mutation in one of the versions of ALA synthase (ALAS 2) which is located on the X-chromosome

Question 71

hemin (lippin)

Answer 71

decreases activity of ALAS1 by causing decrease in synthesis of the enzyme through inhibition of mRNA synthesis and use and by inhibiting mitochondrial import of the enzyme when porphyrin production exceeds the availability of the apoproteins that require it => heme accumulation and is converted to hemin by oxidation of Fe2+ to Fe3++

Question 72

drugs on ALA synthase activity (lippin)

Answer 72

administration of any large number of drugs results in increased hepatic ALAS1 activity drugs metabolized by microsomal cytochrome P450 monooxygenase system in response, synthesis of cytochrome P450 proteins increases => enhanced consumption of heme => decrease in heme concentration in liver => increase in synthesis of ALAS1 and so increase in ALA synthesis

Question 73

porphobilinogen formation (lippin)

Answer 73

when two molecules of ALA are condensed by ALA dehydratase/porphobilinogen synthase extremely sensitive to inhibition by heavy metals because enzyme contains Zn and heavy metals, like Pb, can replace the Zn and inhibit enzyme results in elevation in ALA and anemia