Eicosanoid synthesis
- 1. Eicosanoid synthesis Dr. Radhakrihna G Pillai Department of Life Sciences University of Calicut, India 673 635
- 2. Eicosanoids • The term "eicosanoids" is used as a collective name for molecules derived from 20-carbon fatty acids • Fatty acids of the n-6 family deriving from linoleic acid are the main source of eicosanoids • Arachidonic acid (20:4n-6) being the major precursor • Eicosanoids are synthesized in vivo through several routes • some compounds being formed by more than one mechanism • In the plant kingdom, several potent derivatives from linolenic acid (octadecanoid-derived compounds) are found and have hormone-like functions (phytohormones)
- 3. Lipid number • Lipid numbers. 18:3. ... Lipid numbers take the form C:D, where; – C is the number of carbon atoms in the fatty acid and – D is the number of double bonds in the fatty acid • 18:4 ω-3 or 18:4 n−3 indicates; – an 18-carbon chain – with 4 double bonds and – with the first double bond in the third position from the CH3 end Fig: source https://upload.wikimedia.org/wikipedia/commons/e/e4/Fatty_acid_numbering.png
- 4. Eicosanoid synthesis • Eicosanoids consists of prostaglandins, thromboxanes, leukotrienes and lipoxins • Prostaglandins and thromboxanes are identified as prostanoids • The number of C-C double bonds are used as subscripts with the name of the prostanoids • Majority of biologically active prostaglandins and thromboxanes are referred as series 2 molecules – presence of 2 C-C double bonds • Predominant Leukotrienes are series 4 molecules -4 C-C double bonds • Important series 1 prostaglandins and thromboxanes are also present • Prostaglandins are originally shown to be synthesised in the prostate gland • Thromboxanes from platelets (thrombocyte) • Leucotrenes from leucocytes • Lipoxins are synthesised through lipoxygenase interactions
- 5. Names of Prostaglandins • Prostaglandins are originally shown to be synthesised in the prostate gland • Thromboxanes from platelets and • Leukotrienes from leukocytes • Their names are derived from the place of synthesis https://www.google.co.in/search?q=p latelets&rlz=1C1VFKB_enIN599IN599 &source=lnms&tbm=isch&sa=X&ved= 0ahUKEwjS45uNr53QAhUIvo8KHVv1A JUQ_AUICCgB&biw=1280&bih=621#i mgrc=O0EbkI9EC2Sl8M%3A Source of Fig https://www.google.co.in/search?q=leuk ocytes&rlz=1C1VFKB_enIN599IN599&sou rce=lnms&tbm=isch&sa=X&ved=0ahUKE wjP2a_Mr53QAhXFo48KHeeZAeYQ_AUIC CgB&biw=1280&bih=621#imgrc=NswWA woMbeBGIM%3A
- 6. Eicosanoid synthesis • Two main pathways involved in the biosynthesis of eicosanoids – PG and TX are synthesised by the cyclic pathway – The leukotrenes by the linear pathway • Cyclic pathway initiated by Prostaglanding G/H synthase – This enzyme has 2 activities – COX and peroxidase • Two forms of COX- COX1 an dCOX2 – COX-1 expressed constitutively – gastric mucosa, kidney, platelets and vascular endothelial cells – COX-2 is inducible-in macrophages and monocytes express in response to inflammation • COX-1 and COX-2 catalyse – arachidonic acid to PGG2 and PGH2
- 7. Cyclic pathway • Numerous stimuli- epinephrine, thrombin and bradykinin activates phospholipase A2-(PLA2) • PLA2 hydrolyses arachidonic acid from cell membrane phospholipids • Bradykinin receptor coupled with G-protein activation • Increased intracellular calcium ions • Protein kinase C activated • PKC phosphorylation and Ca2+ ions activate ER membrnane- associated cPLA2 isoform
- 8. Synthesis of prostaglandins • Hydrolysis of arachidonic acid form phosphatidylinositol bisphosphate (PIP2) • Arachidonic acid is converted to PGH2 –action of COX-1 and COX-2 • Prostaglandin is synthesised from PGH2 • Action of different PGE enzymes catalyse the synthesis of different PGs • Thromboxanes are synthesised from PGH2 by thromboxane synthase
- 10. Linear Synthetic pathways • Linear pathway initiated through arachidonate lipoxygenase (LOXs) • 3 forms of LOXs • Arachidonate 5-lipoxygenase (5-LOX), 12-LOX and 15-LOX • 5-LOX help to produce leukotrenes – synthesised by leukocytes, mast cells, lung, spleen, brain and heart • 12-LOX and 15-LOX involved in the synthesis of lipoxins
- 11. Synthesis of Leukotrienes • Numerous stimuli- epinephrine, thrombin, bradykinin activates phospholipase A2-(PLA2) • PLA2 hydrolyses arachidonic acid from cell membrane phospholipids • Bradykinin receptor coupled with G-protein activation • Increased intracellular calcium ions • Protein kinase C activated • PKC phosphorylation and Ca2+ ions activate ER membrnane- associated cPLA2 isoform
- 12. Leukotrienes • Activated cPLA2 isoforms Hydrolyse arachidonic acid from phosphatidylinositol biphosphate (PIP2) • Enzyme 5-LOX + 5-LOX activating protein catalyse the conversion of arachidonic acid to 5- hydroperoxyeicosatetraenoic acid (5-HPETE) • 5-hydroperoxyeicosatetraenoic acid (5-HPETE) is spontaneously reduced to 5- hydroxyeicosatetraenoic acid (5-HETE) • Then to LTA4 • LTA4 unstable – converted to LTC4
- 14. Synthesis of Lipoxins • Synthesised through the concerted actions of 15- LOX on arachidonic acid in epithelial cells, 5-LOX in leukocytes and 12-LOX in platelets • Three pathways – Classic pathway is 5-LOX activity in leukocyte followed by 12-LOX in platelets – 15-LOX in epithelial cells followed by 5-Lox in leukocytes – Action of Aspirin on COX-2 in epithelial or endothelial cells –produce 15-epi-lipoxins (aspiring triggered lipoxins ATLS)
- 16. Porphyrins • Large heterocyclic organic ring structures • Composed of 4 modified pyrrole subunits connected by methine (=CH-) bridges • Heme an example of naturally occurring porphyrin • Heme in biological system consits of Fe2+ ions complexed with 4N of porphyrin molecules Three structurally distinct hemes in human; heme a, heme b and heme c heme is critical for biological functions of several enzymes – eg cytochromes of oxidative phosphorylation Cytochrome P450 family (CYP)
- 17. Heme synthesis • First reaction in mitochondria • Condensation of one succinyl-coA by pyridoxal phosphate – requiring enzyme (vitamin B6) – δaminolevulinic acid synthase (ALAS) – forming δ aminolevulinic acid (5 aminoleuvinic acid) • This is the rate limiting reaction in heme synthesis • ALA is transported to cytosol • ALA dehydratase (porphobilinogen synthetase) dimerises 2molcules of ALA • Forms Porphobilinogen • Head-to-tail condensation of 4 molecules of porphobilinogen -form linear tetrapyrrole intermediate – hydroxymethylbilane
- 18. Heme synthesis • Enzyme for Head-to-tail condensation of 4 molecules of porphobilinogen is porphobilinogen deaminase (PBG deaminase) • Hydroxymethylbilane has two fates • One due to enzymatic action • Other due to non-enzymatic action • Hydroxymethylbilane is enzymatically converted to uroporphyrinogen III • Mediated by the enzyme uroporphyrinogen III synthase • uroporphyrinogen III is decarboxylated by uroporphyrinogen decarboxylase • Forms coproporphyrinogens • Coproporphyrinogen III is most important in heme synthesis • Coproporphyrinogen III transported to the interior of the mitochondria
- 19. Formation of conjugated ring • 2 propionate residues Coproporphyrinogen III are decarboxylated • Protoporphyrinogen IX formed • Catalysed by coproporphyrinogen-III oxidase
- 20. Insertion of Fe2+ • Protoporphyrinogen IX converted to protoporphyrin IX –protophyrinogen IX oxidase • Oxidation reaction requires molecular oxygen • Ring system -Loss of 6protons and 6 electrons – produce a completely conjugated Responsible for the red colour of heme Final reaction in heme synthesis takes place in mitochondria Insertion of Fe2+ into the ring system Enzyme involved is ferrochelatase
- 21. Respiratory burst in phagocytes • Respiratory burst (oxidative burst) -rapid release of reactive oxygen species (superoxide radical and hydrogen peroxide) from different types of cells • Release of these chemicals from immune cells, – e.g., neutrophils and monocytes, as they come into contact with different bacteria or fungi. – They are also released from the ovum of higher animals after the ovum has been fertilized. • Respiratory burst plays an important role in the immune system • Crucial reaction that occurs in phagocytes to degrade internalized particles and bacteria • NADPH oxidase, an enzyme family in the vasculature (in particular, in vascular disease), produces superoxide, which spontaneously recombines with other molecules to produce reactive free radicals
- 22. Phagocytic killing • Oxygen dependent killing • Oxygen independent killing • Oxygen dependent phagocytic killing – Activated phagocytes produce a number of reactive oxygen intermediates and – Nitrogen intermediates • When exposed to certain stimuli- phagocytes (Neutrophils, oesinophils, and macrophages) increase oxygen uptake- upto 50 fold • Oxygen burst
- 23. Superoxide radical • Elimination of invading micro-organisms by neutrophils, monocytes, and macrophages • depends heavily on the generation of reactive oxygen species during the phagocytosis-associated respiratory burst • NADPD oxidase also called respiratory burst oxidase • Present in phagocyte membrane • toxic oxidants are released to the inside and outside of the cell • Catalyse reduction of O2 - by adding electron • 2O2 + NADPH 2O2 - + NADP+ + H+ • 2O2 - + H+ H2O2 - + O2 • the oxidants superoxide anion (O2), hydrogen peroxide (H2O2), hypochlorous acid, and hydroxyl radical, created in this process- • carry the potential to damage the phagocytes themselves as well as other cells at sites of inflammation
- 24. Oxygen-dependent myeloperoxidase- independent intracellular killing • During phagocytosis, glucose is metabolized via the pentose monophosphate shunt, with formation of NADPH • Cytochrome B from the granulocyte-specific granule combines with and activates plasma membrane NADPH oxidase • The activated NADPH oxidase then employs oxygen to oxidize the formed NADPH with resultant production of superoxide anion • A portion of the superoxide anion is converted to H2O2 plus singlet oxygen by superoxide dismutase • Superoxide anion can react with H2O2, resulting in the formation of hydroxyl radical plus more singlet oxygen • Together these reactions produce the toxic oxygen compounds; – superoxide anion (O2-), – H2O2 – singlet oxygen (1O2) and – hydroxyl radical (OH•).
- 25. Oxygen-dependent myeloperoxidase- dependent intracellular killing • Fusion of azurophilic granules with the phagosome causes release of myeloperoxidase into the phagolysosome • azurophilic granules -A large, coarse, blue- purple membrane- bound organelle in progranulocytes, mylocytes and neutrophils, which acts as a reservoir for digestive and hydrolytic enzymes before delivery to a phagosome • Myeloperoxidase utilizes H2O2 and halide ions (usually Cl-) to produce highly toxic hypochlorite • Some hypochlorite spontaneously breaks down to yield singlet oxygen • Together these reactions produce toxic hypochlorite (OCl-) and singlet oxygen (1O2)
- 26. Detoxification reactions Neutrophils and macrophages are able to protect themselves by detoxifying the toxic oxygen intermediates that they generate Granulocyte self-protection is achieved in reactions employing the dismutation of superoxide anion to hydrogen peroxide by superoxide dismutase and The conversion of hydrogen peroxide to water by catalase