Biosynthesis of Lipids V

Suaib Luqman

Objectives

v To understand the biosynthesis of eicosanoids
v How do prostanoids and leukotriene pathway operates
v What are the significance of Bile acid and its synthesis

Concept Map

3. Description

Eicosanoids Biosynthesis

Within the cells eicosanoids are synthesized as and when required without any storage. They originate from the fatty acid that craft both nuclear as well as cell membrane. The oxygenation of fatty acid results in the production of eicosanoids (prostanoids & leukotrienes) with the aid of two important enzymes namely: Cycloxygenase (COX) and Lipoxygenase (LOX).

1. Prostanoid group of eicosanoids are produced by COX.

2. Leukotrienes are constructed by LOX (isoform 5)

The biosynthesis of eicosanoid begins when cell is trigerred by stimulus such as cytokines, growth factors, mechanical trauma etc. followed by the discharge of phospholipase at the cell membrane with an ultimate movement to the nuclear membrane. At nucleus, phospholipids and their esters is hydrolyzed by Phospholipase A2 and diacylglycerol (DAG) by phospholipase C resulting in the release of essential fatty acid (20C). The step of hydrolysis emerges to determine the rate of eicosanoid formation.

The release of fatty acids is due to a type IV cytosolic phospholipase A2, a strategic player. Cells devoid of cytosolic phospholipase A2 generally lack eicosanoid biosynthesis. The cytosolic phospholipase A2 is explicit for phospholipids which contain GPLA, EPA and AA at SN2 position. Captivatingly, cytosolic phospholipase A2 may perhaps helps in the release of lysophospholipid which happen to become platelet activating factor.

The Role of Reactive Oxygen Species and Peroxidation

The fatty acid is symmetrical and the ensuing eicosanoids are chiral. The high stereo-specificity of free fatty acid allows the oxidation following any one of the below mentioned pathways and molecular oxygen is being added to the eicosanoid pathways via COX or LOX activity. The lipid per-oxidation is perilous to the cells, predominantly whilst adjacent to the nucleus and now intricate mechanisms are known which avert superfluous oxidation. Phospholipases, COX and LOX are firmly controlled along with eight other proteins that get activated to synchronize the production of leukotrienes. Most of these leukotrienes subsist in multiple molecular isoforms. The oxidation by LOX or liberates reactive oxygen species (ROS) and the highly reactive peroxides as the primary products in eicosanoid production. The leukotriene A4 be capable of forming adducts with the DNA of tissues. LOX also engenders cellular damage and association of LOX-15 has been found in the pathogenesis of atherosclerosis in murine models. The process of oxidation in eicosanoid production is partitioned which limits the damage caused by peroxides. Carrier proteins, Epoxide hydrolase and Glutathione S transferases are the enzymes responsible for the eicosanoids biosynthesis. The functions of these enzymes are predominantly involved with detoxification process suggesting that eicosanoid signaling might have progress from the cellular ROS detoxification.

Prostanoid Biosynthesis

The prostanoids are produced by the catalytic conversion of the free essential fatty acids by COX via a 2 step process. In the primary step, 2 oxygen molecules are supplemented as 2 peroxide linkages and a carbon ring (5 membered) is counterfeited in the vicinity of the fatty acid chain resulting in the formation of an unstable, short lived intermediary Prostaglandin G (PGG). When single oxygen is sheds from one of the peroxide linkages, formation of Prostaglandin H (PGH) takes place. Additional prostanoids like PGH1, PGH2 and PGH3 is evolved from PGH by the catalytic activity of dissimilar enzymes.

From the above-mentioned pathway, the steps of the biosynthesis are as under.

1. PGH2 synthase/COX/peroxidase converts Arachidonic acid to PGH2.

2. PGD synthase converts PGH2 to Prostaglandin D (PGD).

3. PGE synthase converts PGH2 to Prostaglandin E (PGE which in turn form Prostaglandin F).

4. Prostacyclin synthase converts PGH2 to prostacyclin (PGI2).

5. Thromboxane synthase converts PGH2 to thromboxanes (TXA2)

The PGE, PGI2 and TXA2 are the three prostanoids classes with idiosyncratic rings in the core of the molecule but vary in their structures. The 5C ring PGH compounds (close relative to all) are being linked by 2 oxygen peroxide resulting in single and unsaturated 5C ring prostaglandins. In prostacyclins, the 5C ring is adjacent to another ring with oxygen while in thromboxanes a 6C ring with soliatry oxygen is present. The formation of PGE2 in viral and bacterial infections emerges to be enthused by IL-1 (interleukin-1 and other cytokines).

COX: The Cyclooxygenase Enzyme

Arachidonic acid (ω 6), a polyunsaturated fatty acid gets converted to PGH2 (precursor of the prostanoids) by COX enzymes. COX enzyme encloses two active sites: a cyclooxygenase site, where PGG2 is formed and a heme site with peroxidase activity which is accountable for the diminution of PGG2 to PGH2. A hydrogen atom is abstracted from arachidonic acid by a tyrosine radical engendered by the heme site with peroxidase activity followed by the reaction of 2 oxygen molecules with the arachidonic acid radical capitulating PGG2. Presently, COX1, COX2 and COX3 are the three molecular isoenzymic forms of COX enzymes are known. Numerous tissues articulate anecdotal echelon of COX1 and COX2 whereas COX3 is supposed to be a splice variant of COX1 with frame shift mutation retaining intron one and favored with COX1 variant (COX1v) or COX1b. Both COX1 and COX2 (also known as PGHS1 and PGHS2) act principally in the similar mode, however, selective inhibition can create a distinction in terms of side effects. In most of the mammalian cells, COX1 is present as a constitutive enzyme whereas COX2 is an inducible enzyme undetectable in the majority of normal tissues but plentiful in activated macrophages and inflammation sites of the cells. COX enzymes have recently been shown to be hypersecreted and upregulated in a variety of carcinomas and play a key role in carcionogenesis. Both COX1 and COX2 also oxygenate Eicosa Pentaenoic Acid (EPA, ω 3) and Dihomo-γ-linolenic acid (DGLA, ω 6) to bestow prostanoids of Series 1 and Series 3 (less inflammatory to that of Series 2). EPA and DGLA are competitive inhibitors with arachidonic acid for the COX pathways explaining the fact why borage and fish oil as a dietary source of EPA and DGLA reduce inflammation.

Leukotrienes Biosynthesis

LOX5 catalyses the conversion of arachidonic acid to leukotrienes via incorporation of an oxygen atom at a particular place in the backbone of arachidonic acid. In leucocytes (basophils, eosinophils, mast cells, monocytes and neutrophils), the LOX pathway operates during activation and resulting in the release of arachidonic acid from the phospholipids of the cell membrane by phospholipase A2 and finally bestowed to 5 lipoxygenase (LOX5) by the LOX5 activating protein. The steps are described as under.

1. Conversion of arachidonic acid to 5 hydroperoxyeicosatetraenoic acid (5 HPETE) by using LOX5 and LOX5 activating protein.

2. Spontaneous reduction of 5 HEPETE to 5 hydroxyeicosatetraenoic acid (5 HETE).

3. LOX5 acts yet again on 5 HETE to form leukotriene A4 (LTA4), an unstable epoxide.

4. In monocytes and neutrophils, leukotriene A4 is converted to the dihydroxy acid leukotriene (LTB4), by LTA4 hydrolase. The LTB4 is a potent chemo-attractant for neutrophils performing at BLT1 and BLT2 receptors on the plasma membrane.

5. In eosinophils and mast cells, LTA4 is linked with the tripeptide glutathione to form LTC4 (first cysteinyl leukotrienes) by LTC4 synthase. Extracellularly, LTC4 through the catalytic action of ubiquitous enzymes forms LTD4 and LTE4.

6. At the cell surface receptors (CysLT1 and CysLT2), the action of cysteinyl leukotrienes (LTC4, LTD4, LTE4) results in the contraction of vascular and bronchial muscle, increases blood vessels permeability, augments mucus secretion in the gut and airway to employ leukocytes to the inflammation sites.

7. Both cysteinyl leukotrienes and LTB4 are partially despoiled in neighboring tissues and eventually happen to be inactive metabolites in the liver.

Biosynthesis of Bile acid

Bile acids also referred to as steroid acids are primarily present in the mammalian bile and when it is amalgamated with a cation (e.g. sodium) it is called bile salts. In other words, the protonated (-COOH) and de-protonated or ionized (-COO-) form are referred to as Bile acid and Bile salts respectively. In humans, taurocholic acid and glycocholic acid salts signify about eighty percent of the entire bile salts. Chenodeoxycholic acid and Cholic acid are the two major acids present in bile. In the bile of human intestine, bile acids, deoxycholic acid and lithocholic acid (7α dehydroxylated derivatives), taurine and glycine conjugates are present. The foremost purpose of bile acid is to assist in micelles formations that prop up dietary fat processing and an increase in the bile secretion exhibit increase bile flow.

In liver, bile acids are produced by the oxidation of cholesterol through cytochrome P450. They get stored in the gallbladder (water removed and salts concentrated) in a conjugated form with glycine or taurine or with glucuronide or sulfate. In humans, cholesterol 7α hydroxylase is the rate-limiting step as the addition of a hydroxyl group at position 7 of the steroid nucleus takes place. During the intake of a meal, the gall bladder secretes its content into the intestine where bile acids emulsify the dietary fats. Other functions of bile acids includes: motivating bile flow to remove liver catabolites, cholesterol elimination, emulsification of lipids and fat soluble vitamins to form micelles which can be transported through the lacteal system and help in reducing the bacterial flora present in the biliary tract and small intestine. At intestinal pH, conjugated bile acids are more competent at emulsifying fats as they are more ionized as compared to unconjugated bile acids. In most of the species, the biosynthesis of bile acids is a foremost route of cholesterol metabolism except humans.

About 800mg of cholesterol per day is produced by human body and half of it is used for the synthesis of bile acid. In intestine, approximately 20-30g of bile acids are secreted daily and almost 90% of it gets reabsorbed by the ileum through active transport and recycled by entero-hepatic circulation that budges the bile salts back to the liver and finally to the gallbladder. The process allocates a squat rate of daily synthesis but high secretion to the digestive system. The fat globules are also broken down into tiny droplets by bile and bile from the slaughtered animals is used in soap preparation.

Summary

In this lecture we learnt about:

The Biosynthesis of Eicosanoids: the prostanoids and leukotrienes
The role of COX and LOX
The Biosynthesis of Bile acid

you can view video on Biosynthesis of Lipids V

Weblinks

Books

Lehninger Principles of Biochemistry by David L. Nelson, Albert‎ L. Lehninger, Michael‎ M. Cox. 2008. https://books.google.co.in/books?isbn=071677108X
Eicosanoid Protocols by Elias A. Lianos. 1999. Page 119 https://books.google.co.in/books?isbn=1592592635
The Role of Polyunsaturated Fatty Acid and Eicosanoid by Waddah Katrangi. 2014 https://books.google.co.in/books?id=YGzIoAEACAAJ
The Eicosanoids by Peter Curtis-Prior. 2004. Page 3 https://books.google.co.in/books?isbn=047002061X
Bile acids: chemistry and physiology of bile acids by William T. Beher. 1976. https://books.google.co.in/books?id=qv9rAAAAMAAJ
Bile Acid Biology and Its Therapeutic Implications by G. Paumgartner. 2005 https://books.google.co.in/books?isbn=1402028938