Chap. 10B. Lipids

1. Chap. 10B. Lipids • Storage Lipids • Structural Lipids in Membranes • Lipids as Signals, Cofactors, and Pigments • Working with Lipids Fig. 10-4a. Fat droplets in human adipose tissue cells.

2. Lipids as Signals, Cofactors, and Pigments The two classes of lipids considered in the Chap. 10A file (storage lipids and structural lipids) are major cellular components. Membrane lipids make up 5% to 10% of the dry mass of most cells, and storage lipids can make up more than 80% of the mass of an adipocyte. With some exceptions (phosphatidylinositols and sphingosine derivatives), these lipids play a passive role in the cell. For example, lipid fuels are simply stored until oxidized by enzymes, and membrane lipids mostly form impermeable barriers around cells and cellular compartments. (Phosphatidylinositols and sphingosine derivatives are involved in several signal transduction processes in cells). We now will discuss another group of lipids, present in much smaller amounts, that have active roles in cell physiology as metabolites and messengers.

3. Eicosanoids (I) Eicosanoids (Fig. 10-18) are paracrine hormones, substances that act only on cells near the point of hormone synthesis instead of being transported in the blood to act on cells in distant tissues or organs. These fatty acid derivatives have a variety of effects on vertebrate tissues. They are involved in reproductive function; in the inflammation, fever, and pain associated with injury or disease; in the formation of blood clots and the regulation of blood pressure; in gastric acid secretion; and in various other processes important in human health or disease. All eicosanoids are derived from arachidonic acid [20:4(∆5,8,11,14)], the 20-carbon polyunsaturated fatty acid from which they take their general name (Greek eikosi, “twenty”). There are three classes of eicosanoids: prostaglandins, thromboxanes, and leukotrienes.

4. Eicosanoids (II) Prostaglandins contain a five-carbon ring originating from the chain of arachidonic acid (Fig. 10-18). Their name derives from the prostate gland, the tissue from which they were first isolated. Prostaglandins have a wide array of functions, including elevation of body temperature (fever) and causing inflammation and pain. Thromboxanes have a six-membered ring containing an ether. They are produced by platelets (also called thrombocytes) and act in the formation of blood clots and the reduction of blood flow to the site of a clot. The nonsteroidal antiinflammatory drugs (NSAIDS), such as aspirin and ibuprofen, inhibit the COX enzyme, which catalyzes an early step in the pathway from arachidonate to prostaglandins and thromboxanes. Leukotrienes, first found in leukocytes, contain three conjugated double bonds. Leukotriene D4, derived from leukotriene A4, induces contraction of the smooth muscle lining the airways of the lung. Overproduction of leukotrienes occurs in asthma and anaphylactic shock. The steroid drug prednisone, for example, inhibits the synthesis of prostaglandins, thromboxanes, and leukotrienes by blocking the release of arachidonic acid from membrane lipids by phospholipase A2.

5. Steroid Hormones (I) Steroids are oxidized derivatives of sterols. They have the sterol nucleus but lack the alkyl chain attached to ring D of cholesterol, and they are more polar than cholesterol (Fig. 10-19). Steroid hormones are transported through the bloodstream on protein carriers from their sites of synthesis to target tissues, where they enter cells. On entering cells, they move to the nucleus where they bind to specific receptor proteins that bind DNA and modulate gene expression and thus metabolism. The major groups of steroid hormones are the male and female sex hormones (e.g., testosterone and ß-estradiol), and the adrenal steroids produced by the adrenal cortex (e.g., cortisol and aldosterone). Cortisol mediates the stress response and glucose metabolism, while aldosterone regulates salt excretion by the kidney.

6. Steroid Hormones (II) The synthetic steroids, prednisone and prednisolone have potent antiinflammatory activity. As noted above they inhibit synthesis of prostaglandins, thromboxanes, and leukotrienes. Prednisone and prednisolone mimic the natural antiinflammatory activity of cortisol and are prescribed for asthma and rheumatoid arthritis, among other disorders. Lastly, the plant steroid, brassinolide, is a potent growth regulator that increases the rate of stem elongation and affects the orientation of cellulose microfibrils in the cell wall during growth.

7. Other Plant Signaling Lipids Plants produce thousands of different lipophilic compounds, volatile substances used to attract pollinators, to repel herbivores, to attract organisms that defend the plant against herbivores, and to communicate with other plants. Jasmonate, for example (see Fig. 12-33) derived from linolenic acid [18:3(∆9,12,15)] in membrane lipids, triggers a plant’s defense systems in response to insect-inflicted damage. The methyl ester of jasmonate is responsible for the characteristic fragrance of jasmine oil, which is used in the perfume industry. Many plant volatiles are derived from fatty acids, or from compounds made by the condensation of five-carbon isoprene units (see below). These include geraniol (the characteristic scent of geraniums), ß-pinene (pine trees), limonene (limes), menthol, and carvone (spearmint), to name but a few.

8. Vitamin D The fat-soluble vitamins, A, D, E, and K, are all derived from isoprene units. Vitamins A and D are precursors of hormones. Vitamin D3, also called cholecalciferol, is normally formed in the skin from 7-dehydrocholesterol in a photochemical reaction driven by UV light absorption (Fig. 10-20). Vitamin D3 itself is not biologically active, but is converted by enzymes in the liver and kidney to 1,25-dihydroxyvitamin D3 (calcitriol). Calcitriol is a hormone that regulates calcium uptake in the intestine and calcium levels in kidney and bone. The deficiency of vitamin D leads to defective bone formation and the disease rickets, for which administration of vitamin D produces a dramatic cure. Vitamin D2 (ergocalciferol) is a commercial product formed by UV irradiation of ergosterol from yeast, which resembles vitamin D3. It is further processed to a calcitriol-like active hormone. Vitamin D2 is added to milk and butter as a dietary supplement. Calcitriol regulates gene expressing by interacting with specific nuclear receptor proteins.

9. Vitamin A Vitamin A1 (retinol) derivatives function as a hormone and as the visual pigment of the vertebrate eye (Fig. 10-21). The vitamin A derivative, retinoic acid, is a hormone that regulates gene expression by binding to nuclear receptor proteins. Retinoic acid is required for the development of epithelial tissues, including the skin. It also is the active ingredient in the drug tretinoin (Retin-A) used in the treatment of acne and wrinkled skin. Retinal, another vitamin A derivative, is the pigment that initiates the response of rod and cone cells of the retina to light, producing a neuronal signal to the brain. Good sources of vitamin A are fish liver oils, liver, eggs, whole milk, and butter. In vertebrates, ß-carotene, the pigment that gives carrots, sweet potatoes, and other yellow vegetables their characteristic color, can be enzymatically converted to vitamin A1 (Fig. 10-21). Deficiency of vitamin A leads to dryness of the skin, eyes, and mucous membranes; retarded development and growth; and night blindness, an early symptom commonly used in diagnosing a vitamin A deficiency.

10. Other Isoprenoids (I) Vitamin E is the collective name for a group of closely related lipids called tocopherols, all of which contain a substituted aromatic ring and a long isoprenoid side chain (Fig. 10-22a). As hydrophobic molecules, tocopherols associate with cell membranes, lipids deposits, and lipoproteins in the blood where they react with and destroy oxygen radicals and other free radicals that would otherwise react with and damage unsaturated fatty acids in membrane lipids. Good sources of vitamin E include vegetable oils, eggs, and wheat germ. Laboratory animals fed vitamin E-depleted diets develop scaly skin, muscular weakness and wasting, and sterility. In humans, vitamin E deficiency is rare, and the principal symptom is fragile red blood cells.

11. Other Isoprenoids (II) Vitamin K (Fig. 10-22b) is a cofactor that is required for the synthesis of prothrombin, a blood protein essential in blood coagulation. Vitamin K deficiency slows blood clotting and therefore can be fatal. Vitamin K1 (phylloquinone) is present in green leafy vegetables. A related molecule that is biologically active, vitamin K2 (menaquinone) is formed by bacteria living in the intestine of vertebrates. The drug warfarin (Fig. 10-22c) is a synthetic compound that inhibits the synthesis of active prothrombin by competing with vitamin K for binding to the enzyme that modifies prothrombin. Warfarin is an important anticoagulant drug which is used to treat patients prone to thromboses. It also is used as a rat poison, causing death by internal bleeding.

12. Other Isoprenoids (III) Ubiquinone (also called coenzyme Q) and plastoquinone (Fig. 10-22d,e) are isoprenoids that function as lipophilic electron carriers in oxidation-reduction reactions used for ATP synthesis in mitochondria and chloroplasts, respectively. Both molecules can carry either one or two electrons and either one or two protons (see Fig. 19-3). Dolichols (Fig. 10-22f) carry the sugar units that are added to glycoproteins and glycolipids during their synthesis. Hydrophobic dolichol molecules are anchored to the membrane where these sugar-transfer reactions take place.

13. Natural Pigments Many natural pigments are lipidic conjugated dienes (Fig. 10-23). Conjugated dienes have carbon chains with alternating single and double bonds. Because this structural arrangement allows the delocalization of electrons, the compounds can be excited by low-energy (visible) light, giving them colors that are visible to humans and other animals. Subtle differences in the chemistry of these compounds produce pigments of strikingly different colors. Birds acquire the pigments that color their feathers red or yellow by eating plant materials that contain carotenoid pigments, such as canthaxanthin and zeaxanthin. The differences in pigmentation between male and female birds are the result of differences in intestinal uptake and processing of carotenoids. Like sterols, steroids, dolichols, fat soluble vitamins, ubiquinone, and plastoquinone, these pigments are synthesized from five-carbon isoprene derivatives.

14. Polyketides Polyketides are diverse natural products with potent biological activities. They are lipids and are made via biosynthetic pathways via reactions (Claisen condensations) similar to those used for synthesis of fatty acids. Polyketides are secondary metabolites, compounds that are not central to an organism’s metabolism, but that serve some subsidiary function that gives their producers an advantage in some ecological niche. Some polyketides used in medicine are shown in Fig. 10-24. Erythromycin is an antibiotic, amphotericin B is an antifungal, and lovastatin is an inhibitor of cholesterol synthesis prescribed to decrease one’s risk of cardiovascular disease.

15. Lipid Methods (I) Because lipids are insoluble in water, their extraction and subsequent fractionation require the use of organic solvents and some techniques not commonly used in the purification of water-soluble molecules such as proteins and carbohydrates. In general, complex mixtures of lipids are separated by differences in polarity or solubility in nonpolar solvents. An overview of methods used to isolate and identify lipids is presented in Fig. 10-25. Neutral lipids (triacylglycerols, waxes, pigments, etc.) are readily extracted from tissues with ethyl ether, chloroform, or benzene, solvents that do not permit lipid clustering driven by hydrophobic interactions. Membrane lipids are more effectively extracted by more polar organic solvents, such as ethanol or methanol, which reduce the hydrophobic interactions between lipid molecules while also weakening the hydrogen bonds and electrostatic interactions that bind membrane lipids to membrane proteins. A commonly used extractant is chloroform, methanol, and water (Fig. 10-25a). After extraction, the lipids remain in the denser chloroform phase, while proteins and sugars partition into the upper methanol/water layer.

16. Lipid Methods (II) Major classes of extracted lipids in the chloroform phase may first be separated by thin-layer chromatography, or by adsorption chromatography (Fig. 10-25b). In thin-layer chromatography, lipids are carried up a silica gel-coated plate by a rising solvent front. Less polar lipids travel farther up the plate than do more polar or charged lipids. Lipid bands can be visualized by a number of stains, such as iodine vapor, which binds reversibly to double bonds in unsaturated fatty acids. The region of the silica gel containing the lipids can be scraped from the plates, the lipid eluted in organic solvent, and mass spectrometry or other methods can be used to identify it and its component fatty acids (Fig. 10-26, not covered). Adsorption chromatography on columns of silica gel, through which solvents of increasing polarity are passed, can also be used to fractionate lipids. Closely related lipid species such as phosphatidylcholine and phosphatidylinositol, can be separated by these techniques.

17. Lipid Methods (III) As an alternative to the above methods of analysis, a “shotgun” approach can be used in which a sample of extracted, unfractionated lipids is directly subjected to high-resolution mass spectrometry of different types and under different conditions to determine the total composition of all the lipids: the lipidome (Fig. 10-25c). The lipidome of a cell or tissue changes during differentiation, diseases such as cancer, and during drug treatment.