Lecture 5: Cytochromes P450 Flashcards by Brooke Thornburg

Cytochromes P450

-powerful detox system
-steroid hormone synthesis
-polyunsaturated fatty acid metabolism
-certain proteins in the ER
-proteins in liver metabolize potential toxins

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P450 Detox system

-first line of defense against toxins
-susbstrates are unusual chemicals (drugs, poisons)
-convert compounds to be more easily excreted

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Super family of P450

-dif ones act on molecules with dif structure
-1000s identified in the organism domains

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Plant P450

-need for biosynthesis of pigments and toxins for protection
-used in metabolic efforts for large-scale medicine production

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Nomenclature

Superfamily > family (CYP#)> subfamily (letter)> members (#)

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P450 families

-sequence identity > 40%
-CYP1, CYP2, CYP3, etc

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P450 subfamilies

-sequence identity > 55%
-CYP1A, CYP1B, CYP1C, etc

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P450 individual members

-numbered
-CYP1A1, CYP1A2, CYP1A3

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Human P450

-18 families
-41 subfamilies
-majority CYP1-4
-other families rarely induced

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CYP1-4

-4 inducible (diet drugs etc) human P450 families
-have several members
-metabolize eicosanoid

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Cytochromes P450 enzymatic function

-monooxygenases
-catalyze O from radical, one O to product, one to water
-product more hydrophilic=better substrate for detoxifying
-requires heme iron

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Endogenous P450 substrates

-cholesterol
-steroid hormones
-fatty acids
-higher substrate specificity than exogenous

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P450 substrates

endogenous vs exogenous
-vary in how many substrates each can metabolize

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Exogenous P450 substrates

-drugs
-food additives
-environmental contaminants

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P450 biological functions

-synthesis: of steroid hormones, vit A and D, lipid-like eicosanoid molecules involved in signaling
-metabolism: of fatty acids and eicosannoids
-Detoxification
-Hydroxylation

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Hydroxylation

-increases aqueous solubility of lipid-soluble substrates

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Substrate variety

= reaction variety

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Drug interactions of cytochrome P450

-responsible for >70% of clearance of drugs
-CYP3A4 acts on 30-50% of most drugs
-some reactions harmful: acetaminophen

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CYP3A4A

-liver
-act on 30-50% of drugs (ex erythromycin (antibiotic))
-catalysis of acetaminophen generates highly reactive compound that causes toxicity at high doses

P450 structure

-integral membrane protein with single heme group
-anchored to membrane by N-terminal helix
-well conserved
-conformational changes can occur in binding

P450 heme group

-site of oxidation
-iron coordinated with porphyrin ring and protein
-6th coord site is open or occupied by O2 or other ligands
-on top of heme just above orange sphere

P450 Catalytic cycle

water and iron3
substrate removes water
iron3 to iron 2 via e-
O2 added = oxy-ferrous (one O has -1 charge)
e- transfer (O-1 to O-2) = peroxanion
Compound I

DOG just look at the slide 7

Reactions catalyzed by P450

-aliphatic and aromatic hydroxylation
-epoxidation
-dealkylation
-N-oxidation

-all add oxygen

aliphatic hydroxylation

R-CH2-CH3 –> R-CH2-CH2OH

Aromatic hydroxylation

add OH to ring

Epoxidation

add oxygen to alkene and makes triangle

Dealkylation

R-CH2-NH-CH3 --> R-CH2-NH-CH2-OH --> R-CH2-NH2 + HCHO -get rid of carbon -N or O or S dealkylation

N-oxidation

add OH to NH2

Lipophilic xenobiotics

-drugs, food additive, environment -oxidized by P450 -exogenous compounds

P450 oxidation of xenobiotics

-exogenous compounds -promotes elimination -phase 1 metabolism -phase 2 biosynthesis rxs like linking to glutatione, sulfate, etc -CYP3A4

CYP3A4

-GI tract, liver -responsible for poor bioavailability of some drugs

P450 isoforms of CYP3A4

-less discriminating -variety of lipophlic substrates -multiple sites of oxidation -lower regioselectivity

Metabolism of xenobiotic outcomes

-inactivation (drug metabolism) -activation (prodrug conversion) -formation of highly toxic metabolite (like benzoapyrene from cig burning)

Benzo[a]pyrene

-weak carcinogen -metabolized multiple times to make stronger carcinogen

Steroid hormone pathways

-endogenous substrates -higher P450 specificity -can involve several P450s -one P450 can catalyze multiple steps -SLIDE 15 mech

Induction of P450

decreased drug plasma levels

inhibition of P450

increased drug plasma levels

Drug-drug interactions

-unintended effects can occur due to altering P450 activity -problematic for drugs requiring tight control of plasma levels -vital importance to patient, critical side effects, small therapuetic index

CYP3A4 induction as cause of drug interactions

-upregulation increases rate of metabolism of drugs in liver -Rifampicin -St. John's wort

Rifampicin (induction)

-antituberculosis drug -inducer of CYP3A4 -makes many drugs less effective -increases elimination of warfarin which might increase risk of undertreating anticoagulation

St. John's-wort

-herbal med for depression -induces CYP3A4 and causes multiple adverse drug reactions

CYP3A4 inhibition as a cause of drug interaction

-certain drug inhibits activity (erythromycin/ketoconazole) -patient takes terfenadine for allergies -increased plasma levels that can cause serious cardiac problems

Terfenadine (seldane)

-H1 antihistamine for seasonal allergies -pro drug rapidly metabolized by CYP3A4 into fexofenadine (the active compound) -CYP3A4 inhibition can lead to cardiac problems -avoid risk by using drug replaced by fexofenadine (allegra)

NAPQI

-generated in a small amount from metabolization of acetaminophen by CYP2E1 -normally conjugated with glutathione (GSH) to generate non toxic form

Acetaminophen-induced liver toxicity

-high doses increase NAPQI and can deplete glutathione pool -NAPQI not conjugated = toxic -35% of liver failure cases

CYP2E1

-metabolizes acetaminophen -induced by alcohol = more NAPQI -BUT alcohol is one of its substrates so it can inhibit the metabolism of other drugs -effect depends on time of drinking and taking acetaminophen

Genetic Polymorphisms of P450: CYP2A6

-nicotine metabolism -SLIDE 21